NaPi2B-TARGETED POLYMER ANTIBODY-DRUG CONJUGATES AND METHODS OF USE THEREOF

ABSTRACT

Disclose herein are dosing regimens for targeted NaPi2b antibody-drug conjugates for treating cancer.

RELATED APPLICATIONS

This application is a continuation of U.S. application Ser. No.16/542,642, filed Aug. 16, 2019, now allowed, which claims priority to,and the benefit of, U.S. Provisional Application Nos. 62/719,189, filedAug. 17, 2018; 62/733,380, filed Sep. 19, 2018, 62/808,376, filed Feb.21, 2019 and 62/854,701, filed May 30, 2019, under 35 USC § 119(e). Thecontents of each of these applications are hereby incorporated byreference in their entireties.

INCORPORATION BY REFERENCE OF SEQUENCE LISTING

The contents of the text file named “MRSN-026_C01US_SeqList.txt”, whichwas created on Jun. 28, 2022 and is 20 KB in size, are herebyincorporated by reference in their entirety.

FIELD OF THE INVENTION

This disclosure relates generally to dosing regimens for administeringNaPi2b targeted polymer antibody-drug conjugates for the treatment ofcancer.

BACKGROUND OF THE INVENTION

NaPi2b (SLC34A2, NaPiIIb, Npt2), a multi-transmembrane, sodium-dependentphosphate transporter (Xu et al. Genomics 62:281-284 (1999)), isnormally expressed at the brush border membrane of mammalian smallintestine and participates in the transcellular inorganic phosphate (Pi)absorption, contributing to the maintenance of phosphate homeostasis inthe body. The expression of NaPi2b at the protein level has beendetected in the liver, at the apical surface of epithelial cells ofmammary, salivary glands, and bronchi, and in the lungs, testis, thyroidgland, small intestine, and uterus. Mutations in NaPi2b have beenassociated with clinical syndromes of alveolar and testicular microlithiasis. NaPi2b is highly expressed in non-squamous non-small celllung cancer (NSCLC), non-mucinous ovarian cancer and papillary thyroidcancer. NaPi2b-positive tissue immunoreactivity is present in 61% ofNSCLC, and 92% ovarian cancer specimens.

Ovarian cancer is one of the most common gynecologic malignancies andthe fifth most frequent cause of cancer death in women. The highmortality rate results in part from the frequent diagnosis of ovariancancer at advanced stages and the mortality rate is approximately 65% ofthe incidence rate. Epithelial tumors of ovary comprise 58% of allovarian neoplasms and more than 90% of malignant tumors of ovary.Debulking surgery and platinum-based combination chemotherapy (includingtaxanes) are current treatment modalities; however, the majority ofpatients with relapsed epithelial ovarian cancer eventually succumb tothe disease. There is a need for novel treatment modalities in ovariancancer, including targeted therapies such as immunotherapy withmonoclonal antibodies or cancer vaccine-based approaches.

NSCLC is any type of epithelial lung cancer other than small cell lungcarcinoma (SCLC). NSCLC accounts for about 85% of all lung cancers. As aclass, NSCLCs are relatively insensitive to chemotherapy, compared tosmall cell carcinoma. When possible, they are primarily treated bysurgical resection with curative intent, although chemotherapy isincreasingly being used both pre-operatively (neoadjuvant chemotherapy)and post-operatively (adjuvant chemotherapy). In the metastatic orinoperative setting, chemotherapy and/or immunotherapy is used, althoughthe disease at this stage is largely incurable and survival timesremains short. There is a need for novel treatment modalities in NSCLC,including targeted therapies such as immunotherapy with monoclonalantibodies or cancer vaccine-based approaches.

Accordingly, a need exists for therapies that target the biologicalactivities of NaPi2b.

SUMMARY OF THE INVENTION

In various aspects the invention provides methods of treating a NaPi2bexpressing tumor in a subject by administering to the subject aNaPi2b-targeted fully human or humanized antibody polymer-drugconjugate. The polymer drug conjugate is administered intravenously byan infusion by at a dose of about between 10 mg/m² to 30 mg/m² on thefirst day of treatment and every three weeks thereafter. For example,the dose is about 11.5 mg/m² to about 12.5 mg/m², about 19.5 mg/m²toabout 20.5 mg/m², about 24.5 mg/m²to about 25.5 mg/m², about 29.5mg/m²to about 31.5 mg/m², Preferably the dose is about 12 mg/m² or about30 mg/m²,

In another aspect the invention provides methods of treating a NaPi2bexpressing tumor in a subject by administering to the subject aNaPi2b-targeted fully human or humanized antibody polymer-drug conjugateintravenously by an infusion by at a dose of about between 10 mg/m² to30 mg/m², about between 10 mg/m² to 36 mg/m² or about between 10 mg/m²to 45 mg/m² on the first day of treatment and every four weeksthereafter. For example, the dose is about 11.5 mg/m² to about 12.5mg/m², about 19.5 mg/m²to about 20.5 mg/m², about 24.5 mg/m²to about25.5 mg/m², about 29.5 mg/m² to about 31.5 mg/m², about 32.5 mg/m² toabout 33.5 mg/m², about 35.5 mg/m² to about 36.5 mg/m², about 39.5 mg/m²to about 41.5 mg/m², about 42.5 mg/m² to about 43.5 mg/m², or about 43mg/m² to about 45 mg/m². Preferably the dose is about 30 mg/m² or about33 mg/m² or about 36 mg/m² or about 40 mg/m² or about 43 mg/m².

Preferably, the dose is about 10 mg/m², 15 mg/m², 20 mg/m², 25 mg/m², 30mg/m², 33 mg/m², 36 mg/m², 40 mg/m², 43 mg/m², 43 mg/m², 44 mg/m², 45mg/m² or the dose is about 10 mg/m², 15 mg/m², 20 mg/m², 25 mg/m², 30mg/m², 33 mg/m², 36 mg/m², 40 mg/m² or the dose is about 10 mg/m², 15mg/m², 20 mg/m², 25 mg/m², 30 mg/m², 33 mg/m², 36 mg/m² or the dose isabout 10 mg/m², 15 mg/m², 20 mg/m², 25 mg/m² or 30 mg/m², 33 mg/m² orthe dose is about 10 mg/m², 15 mg/m², 20 mg/m², 25 mg/m² or 30 mg/m², orthe dose is about 10 mg m², 15 mg/m², 20 mg/m² or 25 mg/m² infusedintravenously once every four weeks.

The NaPi2b antibody contains CDRH1 having the amino acid sequenceGYTFTGYNIH (SEQ ID NO: 5); a CDRH2 having the amino acid sequenceAIYPGNGDTSYKQKFRG (SEQ ID NO: 6); a CDRH3 having the amino acid sequenceGETARATFAY (SEQ ID NO: 7); a CDRL1 having the amino acid sequenceSASQDIGNFLN (SEQ ID NO: 8); a CDRL2 having the amino acid sequenceYTSSLYS (SEQ ID NO: 9); and a CDRL3 having the amino acid sequenceQQYSKLPLT (SEQ ID NO: 10);

The polymer-drug conjugate contains Formula A:

wherein:

the polymer comprises poly(1-hydroxymethylethylene hydroxymethyl-formal)(PHF) having a molecular weight ranging from about 5 kDa to about 10kDa;

m is an integer from 20 to 75,

m₁ is an integer from about 5 to about 35,

m₂ is an integer from about 3 to about 10,

m_(3a) is an integer from 0 to about 4,

m_(3b) is an integer from 1 to about 5,

the sum of m, m₁, m₂, m_(3a), and m_(3b) ranges from about 40 to about75, and m₅ is an integer from about 2 to about 5

The subject is human subject. In some aspects the subject is identifiedas having NaPi2b expression as detected by IHC analysis performed on atest cell population obtained from the subject. In other aspects thesubject is identified as having NaPi2b expression as detected by RNAexpression or a gene signature in a sample obtained from the subject.

In some aspects the NaPi2b-expressing tumor is ovarian cancer, non-smallcell lung cancer (NSCLC), papillary thyroid cancer, endometrial cancer,cholangiocarcinoma, papillary renal cell cancer, clear cell renalcancer, breast cancer, kidney cancer, cervical cancer or salivary ductcancer.

In yet another aspect the subject has epithelial ovarian cancer,fallopian tube cancer, primary peritoneal cancer, platinum resistantovarian cancer, non-squamous NSCLC cancer, progressive, radioactiveiodine-refractory, loco-regional recurrent or metastatic diseasepapillary thyroid cancer or epithelial endometrial cancer.

In yet another aspect the subject having epithelial ovarian cancer issubtyped as high-grade ovarian cancer, low-grade serous ovarian canceror clear cell ovarian cancer.

In yet another aspect the subject having ovarian cancer has receivedprior single agent chemotherapy such as, for example, pegylatedliposomal doxorubicin, weekly treatment with paclitaxel topotecangemcitabine, PARP inhibitor and the like.

In yet another aspect the subject having ovarian cancer has received nomore than 3 lines of prior lines of therapy such as, for example,including but not limited to, chemotherapy combination, such as, forexample, carboplatin plus paclitaxel, pegylated liposomal doxorubicin,weekly treatment with paclitaxel, docetaxel, topotecan, gemcitabine,PARP inhibitor and the like. In a further aspect the subject havingNSCLC cancer is subtyped as adenocarcinoma.

In another aspect the subject has NSCLC and has received priortreatment, such as for example, with a platinum-based chemotherapy(cisplatin or carboplatin) and a PD-1 or PD-L1 monoclonal antibody. Inanother aspect the subject has NSCLC and has received prior treatmentwith carboplatin/paclitaxel, abraxane nab-paclitaxel, docetaxel,premetrexed, gemcitabine or a combination of docetaxel and ramucirumab.

In another aspect the subject has NSCLC and has not received additionalprior treatment with a cytotoxic agent or has not receivedimmunotherapy. In another aspect the subject having NSCLC has documentedintolerance or disease progression with known oncogenic mutations forwhich there are approved therapies (e.g. ALK translocation, EGFRmutation).

In a further aspect the subject having NSCLC cancer is treated with apolymer-drug conjugate of Formula A and a PD-1 or PD-L1 monoclonalantibody, such as, for example, nivolumab, pembrolizumab, atezolizumabor avelumab

In another aspect the subject has papillary thyroid cancer withresistance or intolerance to prior kinase inhibitor therapy or hasreceived prior treatment for low-grade, hormone receptor-positiveendometroid adenocarcinoma.

In another aspect the endometrial cancer is not a stromal tumor or acarcinosarcoma.

In another aspect the subject has endometrial cancer and has receivedprior treatment with a carboplatin/paclitaxel or a similar regimen.

In another aspect the subject has papillary renal cell cancer or clearcell renal cancer that has a predominantly papillary growth pattern. Inone aspect the subject has a histologic diagnosis of salivary ductcancer has progressed after standard systemic therapy.

In yet another aspect, the subject is refractory to chemotherapy,including standard, front-line chemotherapeutic agents.

In a further aspect the subject is treated with a polymer-drug conjugateof Formula A in combination with a PARP inhibitor, such as, for example,olaparib, niraparib, rucaparib, talazoparib, and the like; a PD1/PDL-1inhibitor, such as, for example, nivolumab, pembrolizumab, atezolizumab,avelumab, and the like; chemotherapy, such as, for example, carboplatin,cisplatin, oxaliplatin, doxil, cyclophosphamide, gemcitabine, topotecan,premetrexe, and the like; a VEGF inhibitor, such as, for example,bevacizumab, ramucirumab, and the like; a tyrosine kinase inhibitor,such as, for example, gefitinib, afatinib, erlotinib, dacomitinib,osimertinib, pazopanib, and the like; an ALK inhibitor, such as, forexample, alectinib, crizotinib, certinib, brigatinib, and the like; or aBRAF inhibitor, such as, for example, dabrafenib, trametinib, and thelike.

In another aspect, the subject is treated with a polymer-drug conjugateof Formula A in combination with pembrolizumab, carboplatin, doxil,bevacizumab or a PARP inhibitor. Unless otherwise defined, all technicaland scientific terms used herein have the same meaning as commonlyunderstood by one of ordinary skill in the art to which this inventionbelongs. In the specification, the singular forms also include theplural unless the context clearly dictates otherwise. Although methodsand materials similar or equivalent to those described herein can beused in the practice or testing of the present invention, suitablemethods and materials are described below. In the case of conflict, thepresent specification, including definitions, will control. In addition,the materials, methods and examples are illustrative only and are notintended to be limiting.

Other features and advantages of the invention will be apparent from thefollowing detailed description and claims.

BRIEF DESCRIPTION OF THE FIGURES

FIG. 1 is a swimmers plot that summarizes details for the time on studyfor all patients though DL6, 21 day cycle, (n=19) and four patient dosedpatients in the 28-day cycle (n=4).

FIG. 2 is a “waterfall plot” that shows the best response by RECISTcriteria for the patients in DL1 to DL5 on the 21-day treatment cyclewho have had at least one scan to date.

FIG. 3 shows the best overall response for patients (n=11) on the 21-daytreatment cycle.

FIG. 3 shows the best overall response for patients (n=11) on the 21-daytreatment cycle.

FIG. 4 is a “waterfall plot” that shows the best response by RECISTcriteria for the patients dosed in the 21-day cycle and 28-day cycle andwho have had at least one scan to date.

FIG. 5 is a “swimmers plot” that summarizes the details for the time onstudy for patients though DL6, 21-day cycle (n=20) and DL 4A and DL 5Apatients dosed in the 28-day cycle (n=16).

FIG. 6 shows the H-score obtained from NaPi2b protein expression by IHCfrom 34 patient tumor samples. The y axis shows the NaPi2b H-score andthe x axis shows the tumor type.

FIG. 7 is a “swimmers plot” that summarizes the details for the time onstudy for patients though DL6, 21-day cycle (n=20) and. DL 4A and DL 5Apatients dosed in the 28-day cycle (n=17).

FIG. 8 is a “swimmers plot” that summarizes the details for the time onstudy for patients across 7 dose levels (2 to 40 mg/m²), 21-day cycleand 3 dose levels (20 to 36 mg/m²), 28-day cycle.

DETAILED DESCRIPTION

The present disclosure provides methods of treating NaPi2b expressingcancer by administering a NaPi2b-targeted polymer antibody-drugconjugate that specifically bind to the extracellular region of SLC34A2.Specifically, the invention provides dosing regimens for XMT-1536 in thetreatment of NaPi2b expressing cancers. XMT-1536 is comprised of about10-15 molecules of auristatin F-hydroxypropyl amide (AF HPA) conjugatedto a cysteine moiety of a NaPi2b monoclonal antibody (XMT-1535) via apoly(1-hydroxymethylethylene hydroxymethylformal) (PHF) scaffold.

Patients with NaPi2b-expressing ovarian cancer, NSCLC, papillary thyroidcancer, endometrial cancer, papillary renal cell cancer or salivary ductcancer were intravenously administered XMT-1536 every three weeks in adose escalation study. The disease control rates was 67% for patientstreated with at least 12 mg/m² XMT-1536 as per RECIST, version 1.1.XMT-1536 has been well tolerated at the maximum doses administered (40mg/m²). Accordingly, the invention features methods of treating NaPi2bexpressing tumors by administering to a subject, i.e., human, aninfusion dose of about between 10 to 45 mg/m². Tumors include ovariancancer, non-small cell lung cancer (NSCLC), papillary thyroid cancer,endometrial cancer, cholangiocarcinoma, papillary renal cell cancer,clear cell renal cancer, breast cancer, kidney cancer, cervical canceror salivary duct cancer. The tumor is a primary tumor or a metastatictumor.

Patients with NaPi2b-expressing ovarian cancer, NSCLC, papillary thyroidcancer, endometrial cancer, papillary renal cell cancer or salivary ductcancer were intravenously administered XMT-1536 every four weeks in adose escalation study with an infusion dose of about between 20 to 45mg/m² i.e. 20 mg/m², 25 mg/m², 30 mg/m², 33 mg/m², 36 mg/m², 40 mg/m²,43 mg/m², 44 mg/m², or 45 mg/m². The subject may or may not havereceived previous treatment for the cancer. For example, the subject hasreceived platinum-based chemotherapy, PD-1 or PD-L1 regimens, orpaclitaxel.

In some aspects the subject has been identified as having NaPi2bexpression. NaPi2b expression is detected by methods known in the art.For example, by immunohistochemistry (IHC) analysis, fluorescent in situhybridization (FISH) assay or RNA expression analysis.

NaPi2b Antibodies

The NaPi2b antibodies suitable for the methods of the disclosurespecifically bind specific binding to the extracellular region ofSLC34A2. The disclosure further provides NaPi2b-targeted monoclonalantibodies that specifically recognizes NaPi2b, also known assodium-dependent phosphate transport protein 2B. The NaPi2b antibodiesused in the conjugates disclosed herein are capable of and useful inmodulating, e.g., blocking, inhibiting, reducing, antagonizing,neutralizing or otherwise interfering with at least one biologicalactivity of NaPi2b. Antibodies disclosed herein also include antibodiesthat bind soluble NaPi2b. The NaPi2b antibodies specifically bind to anepitope on an extracellular domain (ECD) of the human NaPi2b. Theseantibodies are collectively referred to herein as “NaPi2b” antibodies.

The NaPi2b antibody-drug conjugates provided herein include antibodiesthat bind to a NaPi2b epitope with an equilibrium dissociation constant(K_(d) or K_(D)) of ≤1 μM, e.g., ≤100 nM, preferably ≤10 nM, and morepreferably ≤1 nM. For example, the NaPi2b antibodies used in theantibody-drug conjugates disclosed herein exhibit a K_(d) in the rangeapproximately between ≤1 nM to about 1 pM.

The NaPi2b antibody-drug conjugates provided herein can includeantibodies that serve to modulate, block, inhibit, reduce, antagonize,neutralize or otherwise interfere with the functional activity ofNaPi2b. Functional activities of NaPi2b include for example,participating in the transcellular inorganic phosphate (Pi) absorption,thereby contributing to the maintenance of phosphate homeostasis in thebody. For example, the NaPi2b antibodies completely or partially inhibitNaPi2b functional activity by partially or completely modulating,blocking, inhibiting, reducing antagonizing, neutralizing, or otherwiseinterfering with transcellular inorganic phosphate absorption.Transcellular inorganic phosphate absorption activity is assessed usingany art-recognized method for detecting transcellular inorganicphosphate absorption activity, including, but not limited to detectinglevels of transcellular inorganic phosphate absorption in the presenceand absence of an anti-NaPi2b antibody disclosed herein.

The NaPi2b antibodies are considered to completely modulate, block,inhibit, reduce, antagonize, neutralize or otherwise interfere withNaPi2b functional activity when the level of NaPi2b functional activityin the presence of the NaPi2b antibody is decreased by at least 95%,e.g., by 96%, 97%, 98%, 99% or 100% as compared to the level of NaPi2bfunctional activity in the absence of binding with a NaPi2b antibodydescribed herein. The NaPi2b antibodies are considered to partiallymodulate, block, inhibit, reduce, antagonize, neutralize or otherwiseinterfere with NaPi2b functional activity when the level of NaPi2bactivity in the presence of the NaPi2b antibody is decreased by lessthan 95%, e.g., 10%, 20%, 25%, 30%, 40%, 50%, 60%, 75%, 80%, 85% or 90%as compared to the level of NaPi2b activity in the absence of bindingwith a NaPi2b antibody described herein.

Exemplary antibodies disclosed herein include, the XMT-1535 antibody.These antibodies show specificity for human NaPi2b and they have beenshown to inhibit NaPi2b activity.

NaPi2b human or humanized monoclonal antibody, XMT-1535, includes aheavy chain (HC), heavy chain variable region (VH), light chain (LC),and a light chain variable region (VL), as shown in the amino acid andcorresponding nucleic acid sequences presented below. The variable heavychain region and variable light chain region for each antibody areshaded in the amino acid sequences below. The complementaritydetermining regions (CDRs) of the heavy chain and the light chain areunderlined in the amino acid sequences presented below. The amino acidsencompassing the complementarity determining regions (CDRs) for theXMT-1535 antibody are as defined by E. A. Kabat et al. (See Kabat, E.A., et al., Sequences of Protein of immunological interest, FifthEdition, US Department of Health and Human Services, US GovernmentPrinting Office (1991)) and are disclosed in U.S. Pat. No. 8,603,474.

>XMT-1535 Heavy Chain Amino Acid Sequence (Heavy chain variableregion (SEQ ID NO: 3) (Italicized) + IgG1 Heavy chain constantregion (SEQ ID NO: 11)) (SEQ ID NO: 1)

ASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKRVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSREEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPG* (SEQ ID NO: 5)CDRH1: GYTFTGYNIH (SEQ ID NO: 6) CDRH2: AIYPGNGDTSYKQKFRG (SEQ ID NO: 7)CDRH3: GETARATFAY >XMT-1535 Heavy chain variable region nucleic acid sequence(SEQ ID NO: 13)CAAGTTCAGCTGGTTCAGICTGGCGCCGAGGTTGTGAAACCTGGCGCCTCTGTGAAGATGAGCTGCAAGGCCAGCGGCTACACCTTCACCGGCTACAACATCCACTGGGTCAAGCAGGCCCCTGGACAGGGACTCGAATGGATCGGAGCCATCTATCCCGGCAACGGCGACACCAGCTACAAGCAGAAGTTCCGGGGCAGAGCCACACTGACCGCCGATACAAGCACCAGCACCGTGTACATGGAACTGAGCAGCCTGAGAAGCGAGGACAGCGCCGTGTACTATTGCGCCAGAGGCGAAACAGCCAGAGCCACCTTTGCCTATTGGGGCCAGGGAACCCTGGTCACCGTTAGCTCT >XMT-1535 Light Chain Amino Acid Sequence (Light chain variableregion (SEQ ID NO: 4) (Italicized) + Light chain constant region(SEQ ID NO: 12)) (SEQ ID NO: 2)

RTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC (SEQ ID NO: 8) CDRL1: SASQDIGNFLN (SEQ ID NO: 9)CDRL2: YTSSLYS (SEQ ID NO: 10)CDRL3: QQYSKLPLT >XMT-1535 Light chain variable region nucleic acid sequence(SEQ ID NO: 14)GATATTCAGATGACACAGAGCCCCAGCAGCCTGTCTGCCTCTGTGGGAGACAGAGTGACCATCACCTGTAGCGCCAGCCAGGATATCGGCAACTTCCTGAACTGGTATCAGCAGAAACCCGGCAAGACCGTGAAGGTGCTGATCTACTACACCTCCAGCCTGTACAGCGGCGTGCCCAGCAGATTTTCTGGCAGCGGCTCTGGCACCGACTACACCCTGACCATATCTAGCCTGCAGCCTGAGGACTTCGCCACCTACTACTGCCAGCAGTACAGCAAGCTGCCCCTGACATTTGGCCAGGGCACCAAGCTGGAACTGAAG

Also included in the disclosure are antibodies that bind to the sameepitope or cross compete for binding to the same epitope as theantibodies described herein. For example, antibodies disclosed hereinspecifically bind to NaPi2b, wherein the antibody binds to an epitopethat includes one or more amino acid residues on human NaPi2b (e.g.,GenBank Accession No. 095436.3).

Antibodies disclosed herein specifically bind to an epitope on thefull-length human NaPi2b comprising the amino acid sequence:

(SEQ ID NO: 15) 1 MAPWPELGDA QPNPDKYLEG AAGQQPTAPD KSKETNKTDN TEAPVTKIEL51 LPSYSTATLI DEPTEVDDPW NLPTLQDSGI KWSERDTKGK ILCFFQGIGR 101LILLLGFLYF FVCSLDILSS AFQLVGGKMA GQFFSNSSIM SNPLLGLVIG 151VLVTVLVQSS STSTSIVVSM VSSSLLTVRA AIPIIMGANI GTSITNTIVA 201LMQVGDRSEF RRAFAGATVH DFFNWLSVLV LLPVEVATHY LEIITQLIVE 251SFHFKNGEDA PDLLKVITKP FTKLIVQLDK KVISQIAMND EKAKNKSLVK 301IWCKTFTNKT QINVTVPSTA NCTSPSLCWT DGIQNWTMKN VTYKENIAKC 351QHIFVNFHLP DLAVGTILLI LSLLVLCGCL IMIVKILGSV LKGQVATVIK 401KTINTDFPFP FAWLTGYLAI LVGAGMTFIV QSSSVFTSAL TPLIGIGVIT 451IERAYPLTLG SNIGTTTTAI LAALASPGNA LRSSLQIALC HFFFNISGIL 501LWYPIPFTRL PIRMAKGLGN ISAKYRWFAV FYLIIFFFLI PLTVFGLSLA 551GWRVLVGVGV PVVFIIILVL CLRLLQSRCP RVLPKKLQNW NFLPLWMRSL 601KPWDAVVSKF TGCFQMRCCC CCRVCCRACC LLCDCPKCCR CSKCCEDLEE 651AQEGQDVPVK APETFDNITI SREAQGEVPA SDSKTECTAL

Antibodies disclosed herein specifically bind to an epitope on anextracellular domain (ECD) of the human NaPi2b.

Those skilled in the art will recognize that it is possible todetermine, without undue experimentation, if a monoclonal antibody hasthe same specificity as a monoclonal antibody disclosed herein (e.g.,XMT-1535, 10H1.11.4B) by ascertaining whether the former prevents thelatter from binding to a natural binding partner or other molecule knownto be associated with NaPi2b. If the monoclonal antibody being testedcompetes with the monoclonal antibody disclosed herein, as shown by adecrease in binding by the monoclonal antibody disclosed herein, thenthe two monoclonal antibodies bind to the same, or a closely related,epitope.

An alternative method for determining whether a monoclonal antibody hasthe specificity of monoclonal antibody disclosed herein is topre-incubate the monoclonal antibody disclosed herein with solubleNaPi2b (with which it is normally reactive), and then add the monoclonalantibody being tested to determine if the monoclonal antibody beingtested is inhibited in its ability to bind NaPi2b. If the monoclonalantibody being tested is inhibited then, in all likelihood, it has thesame, or functionally equivalent, epitopic specificity as the monoclonalantibody disclosed herein.

Screening of monoclonal antibodies disclosed herein, can also be carriedout, e.g., by measuring NaPi2b-mediated activity, and determiningwhether the test monoclonal antibody is able to modulate, block,inhibit, reduce, antagonize, neutralize or otherwise interfere withNaPi2b activity.

The antibodies disclosed herein contain a heavy chain variable regionhaving an amino acid sequence at least 90%, 91%, 92%, 93%, 94%, 95%,96%, 97% 98%, 99% or more identical to a sequence selected from thegroup consisting of SEQ ID NOs: 3 and a light chain variable regionhaving an amino acid sequence at least 90%, 91%, 92%, 93%, 94%, 95%,96%, 97% 98%, 99% or more identical to a sequence selected from thegroup consisting of SEQ ID NOs: 4.

In some embodiments, the antibodies disclosed herein contain a heavychain amino acid sequence at least 90%, 91%, 92%, 93%, 94%, 95%, 96%,97% 98%, 99% or more identical to the amino acid sequence of SEQ ID NO:1 and a light chain amino acid sequence at least 90%, 91%, 92%, 93%,94%, 95%, 96%, 97% 98%, 99% or more identical to the amino acid sequenceof SEQ ID NO: 2.

The antibodies disclosed herein contain a heavy chain variable regionhaving an amino acid sequence at least 85%, 86%, 87% 88%, 89%, 90%, 91%,92%, 93%, 94%, 95%, 96%, 97% 98%, 99% or more identical to a sequenceselected from the group consisting of SEQ ID NOs: 3 and a light chainvariable region having an amino acid sequence at least 85%, 86%, 87%88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97% 98%, 99% or moreidentical to a sequence selected from the group consisting of SEQ IDNOs: 4.

In some embodiments, the antibodies disclosed herein contain a heavychain amino acid sequence at least 85%, 86%, 87% 88%, 89%, 90%, 91%,92%, 93%, 94%, 95%, 96%, 97% 98%, 99% or more identical to the aminoacid sequence of SEQ ID NO: 1 and a light chain amino acid sequence atleast 85%, 86%, 87% 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%98%, 99% or more identical to the amino acid sequence of SEQ ID NO: 2.

In some embodiments, the antibodies disclosed herein contain the heavychain variable region amino acid sequence of SEQ ID NO: 3 and the lightchain variable region amino acid sequence of SEQ ID NO: 4.

In some embodiments, the antibodies disclosed herein contain the heavychain amino acid sequence of SEQ ID NO: 1 and the light chain amino acidsequence of SEQ ID NO: 2.

In some embodiments, the antibodies disclosed herein contain the CDRH1amino acid sequence of SEQ ID NO: 5, the CDRH2 amino acid sequence ofSEQ ID NO: 6, the CDRH3 amino acid sequence of SEQ ID NO: 7, the CDRL1amino acid sequence of SEQ ID NO: 8, the CDRL2 amino acid sequence ofSEQ ID NO: 9, and the CDRL3 amino acid sequence of SEQ ID NO: 10.

In some embodiments, the antibodies disclosed herein that contains theamino acid sequence at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97% 98%,99% or more identical to the amino acid sequence GYTFTGYNIH (SEQ ID NO:5); a CDRH2 that contains the amino acid sequence at least 90%, 91%,92%, 93%, 94%, 95%, 96%, 97% 98%, 99% or more identical to the aminoacid sequence AIYPGNGDTSYKQKFRG (SEQ ID NO: 6); a CDRH3 that containsthe amino acid sequence at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%98%, 99% or more identical to the amino acid sequence GETARATFAY (SEQ IDNO: 7); a CDRL1 that contains the amino acid sequence at least 90%, 91%,92%, 93%, 94%, 95%, 96%, 97% 98%, 99% or more identical to the aminoacid sequence SASQDIGNFLN (SEQ ID NO: 8); a CDRL2 that contains theamino acid sequence at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97% 98%,99% or more identical to the amino acid sequence YTSSLYS (SEQ ID NO: 9);and a CDRL3 that contains the amino acid sequence at least 90%, 91%,92%, 93%, 94%, 95%, 96%, 97% 98%, 99% or more identical to the aminoacid sequence QQYSKLPLT (SEQ ID NO: 10).

In some embodiments, the antibodies disclosed herein that contains theamino acid sequence at least 85%, 86%, 87% 88%, 89%, 90%, 91%, 92%, 93%,94%, 95%, 96%, 97% 98%, 99% or more identical to the amino acid sequenceGYTFTGYNIH (SEQ ID NO: 5); a CDRH2 that contains the amino acid sequenceat least 85%, 86%, 87% 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%98%, 99% or more identical to the amino acid sequence AIYPGNGDTSYKQKFRG(SEQ ID NO: 6); a CDRH3 that contains the amino acid sequence at least85%, 86%, 87% 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97% 98%, 99%or more identical to the amino acid sequence GETARATFAY (SEQ ID NO: 7);a CDRL1 that contains the amino acid sequence at 85%, 86%, 87% 88%, 89%,90%, 91%, 92%, 93%, 94%, 95%, 96%, 97% 98%, 99% or more identical to theamino acid sequence SASQDIGNFLN (SEQ ID NO: 8); a CDRL2 that containsthe amino acid sequence at least 85%, 86%, 87% 88%, 89%, 90%, 91%, 92%,93%, 94%, 95%, 96%, 97% 98%, 99% or more identical to the amino acidsequence YTSSLYS (SEQ ID NO: 9); and a CDRL3 that contains the aminoacid sequence at least 85%, 86%, 87% 88%, 89%, 90%, 91%, 92%, 93%, 94%,95%, 96%, 97% 98%, 99% or more identical to the amino acid sequenceQQYSKLPLT (SEQ ID NO: 10).

In certain embodiments, the antibodies disclosed herein include one ormore conservative amino acid substitutions in a variable domain sequencesuch as 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, or moreconservative substitutions in a variable domain sequence. In someembodiments, these conservative amino acid substitutions are in a CDRregion, e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, or moreconservative substitutions are made cumulatively across all CDRs and insome particular embodiments, up to 1, 2, 3, or 4 conservative amino acidsubstitutions may be present in each CDR sequence, e.g., SEQ ID NOs:5-10.

Those skilled in the art will recognize that it is possible todetermine, without undue experimentation, if a monoclonal antibody hasthe same specificity as a monoclonal antibody XMT-1535, by ascertainingwhether the former prevents the latter from binding to a natural bindingpartner or other molecule known to be associated with NaPi2b. If themonoclonal antibody being tested competes with the monoclonal antibodydisclosed herein, as shown by a decrease in binding by the monoclonalantibody disclosed herein, then the two monoclonal antibodies bind tothe same, or a closely related, epitope.

An alternative method for determining whether a monoclonal antibody hasthe specificity of monoclonal antibody disclosed herein is topre-incubate the monoclonal antibody disclosed herein with solubleNaPi2b (with which it is normally reactive), and then add the monoclonalantibody being tested to determine if the monoclonal antibody beingtested is inhibited in its ability to bind NaPi2b. If the monoclonalantibody being tested is inhibited then, in all likelihood, it has thesame, or functionally equivalent, epitopic specificity as the monoclonalantibody disclosed herein.

Screening of monoclonal antibodies disclosed herein, can be also carriedout, e.g., by measuring NaPi2b -mediated activity, and determiningwhether the test monoclonal antibody is able to modulate, block,inhibit, reduce, antagonize, neutralize or otherwise interfere withNaPi2b activity.

The NaPi2b antibodies suitable for use in the methods disclosed hereincan be generated and purified by well-known techniques e.g., WO2009/097128, WO 2017/160754, and US 16/136,706, each of which isincorporated herein in its entirety by reference.

NaPi2b Targeted Polymer Antibody Drug Conjugates

The invention pertains to therapies involving immunoconjugatescomprising an antibody conjugated to a cytotoxic agent such as a toxin(e.g., an enzymatically active toxin of bacterial, fungal, plant, oranimal origin, or fragments thereof), via a polymer scaffold.

The conjugate described herein includes a NaPi2b antibody connected toone or more AF-HPA-carrying polymeric scaffolds independently comprisingpoly(1-hydroxymethylethylene hydroxymethyl-formal) (PHF) having amolecular weight ranging from about 5 kDa to about 10 kDa. TheAF-HPA-carrying polymeric scaffold is conjugated to the NaPi2b targetedantibody via the NaPi2b cysteine residues.

Specifically, the NaPi2b targeted polymer antibody-drug conjugate isXMT-1536 and has the Formula (A):

wherein:

the polymer comprises poly(1-hydroxymethylethylene hydroxymethyl-formal)(PHF) having a molecular weight ranging from about 5 kDa to about 10kDa;

m is an integer from 20 to 75,

m₁is an integer from about 5 to about 35,

m₂is an integer from about 3 to about 10,

m_(3a) is an integer from 0 to about 4,

m_(3b) is an integer from 1 to about 5,

the sum of m, m₁, m₂, m_(3a), and m_(3b), ranges from about 40 to about75,

m₅ is an integer from about 2 to about 5, and NaPi2b is the fully humanor humanized NaPi2b antibody XMT1535 described herein.

In some embodiments, m is an integer from about 30 to about 75.

In some embodiments, m is an integer from about 30 to about 40.

In some embodiments, m₁ is an integer from about 10 to about 20.

In some embodiments, m₁ is an integer from about 10 to about 12.

In some embodiments, m₂ is an integer from about 3 to about 5.

In some embodiments, m_(3a) is an integer from 0 to about 1.

In some embodiments, m_(3b) is an integer from 2 to about 4

In some embodiments, m₅ is an integer from about 3 to about 4.

In some embodiments the NaPi2b targeted polymer antibody-drug conjugatecomprises 10-15 molecules of AF-HPA.

In some embodiments, the PHF has a molecular weight ranging from about 6kDa to about 8 kDa.

In some embodiments, the PHF has a molecular weight ranging from about 6kDa to about 7 kDa.

In certain embodiments, the NaPi2b targeted polymer antibody-drugconjugate Formula (A) is of Formula (B), wherein the polymer is PHF thathas a molecular weight ranging from about 5 kDa to about 10 kDa:

wherein:

m is an integer from 30 to about 35,

m₁ is an integer from 8 to about 10,

m₂ is an integer from 2 to about 5,

m_(3a) is an integer from 0 to about 1,

m_(3b) is an integer from 1 to about 2,

the sum of m_(3a) and m_(3b) ranges from 1 and about 4, and

the ratio between the PHF and the antibody is about 3 to about 5.

The NaPi2b targeted polymer antibody-drug conjugates, (i.e., XMT-1536)suitable for use in the methods disclosed herein can be generated andpurified by well-known techniques e.g., WO 2009/097128, WO 2017/160754,PCT/US18/38988 and U.S. Ser. No. 16/136,706, each of which isincorporated herein in its entirety by reference.

Dosage and Administration

The cancer therapy provided herein, containing a NaPi2b -targetedpolymer antibody-drug conjugate, is administered in an amount sufficientto exert a therapeutically useful effect. Typically, the active agentsare administered in an amount that does not result in undesirable sideeffects of the patient being treated, or that minimizes or reduces theobserved side effects. NaPi2b expressing cancers include for example, ofovarian cancer, non-small cell lung cancer (NSCLC), papillary thyroidcancer, endometrial cancer, cholangiocarcinoma, papillary renal cellcancer, clear cell renal cancer, breast cancer, kidney cancer, cervicalcancer and salivary duct cancer.

It is within the level of one of skill in the art to determine theprecise amounts of active agents, including NaPi2b-targeted polymerantibody-drug conjugates to be administered to a subject. For example,such agents and uses for treating cancers and solid tumors, arewell-known in the art. Thus, dosages of such agents can be chosen basedon standard dosing regimens for that agent under a given route ofadministration.

It is understood that the precise dosage and duration of treatment is afunction of the tissue or tumor being treated and may be determinedempirically using known testing protocols or by extrapolation from invivo or in vitro test data and/or can be determined from known dosingregimens of the particular agent. It is to be noted that concentrationsand dosage values may also vary with the age of the individual treated,the weight of the individual, the route of administration and/or theextent or severity of the disease and other factors that are within thelevel of a skilled medical practitioner to consider. Generally, dosageregimens are chosen to limit toxicity. It should be noted that theattending physician would know how to and when to terminate, interruptor adjust therapy to lower dosage due to toxicity, or bone marrow, liveror kidney or other tissue dysfunctions. Conversely, the attendingphysician would also know how to and when to adjust treatment to higherlevels if the clinical response is not adequate (precluding toxic sideeffects). It is to be further understood that for any particularsubject, specific dosage regimens should be adjusted over time accordingto the individual need and the professional judgment of the personadministering or supervising the administration of the formulations, andthat the concentration ranges set forth herein are exemplary only andare not intended to limit the scope thereof.

For example, the NaPi2b-targeted polymer antibody-drug conjugate, isadministered in a therapeutically effective amount to decrease the tumorvolume.

The amount of a NaPi2b-targeted polymer antibody-drug conjugate isadministered for the treatment of a disease or condition, for example acancer or solid tumor can be determined by standard clinical techniques.In addition, in vitro assays and animal models can be employed to helpidentify optimal dosage ranges. The precise dosage, which can bedetermined empirically, can depend on the route of administration, thetype of disease to be treated and the seriousness of the disease.

The conjugates provided herein are administered intravenously. Forintravenous administration, the conjugate can be administered by push orbolus, by infusion, or via a combination thereof. The infusion time canbe about 1 minute to three hours, such as about 1 minute to about twohours, or about 1 minute to about 60 minutes, or at least 10 minutes, 40minutes, or 60 minutes.

The dosage amount is between about 10 mg/m² to 30 mg/m². For example,the dosage is between about 11.5 mg/m² to about 12.5 mg/m².Alternatively, the dosage is between about 19.5 mg/m²to about 20.5mg/m², between about 24.5 mg/m²to about 25.5 mg/m², between about 29.5mg/m² to about 31.5 mg/m². In some embodiments, the dosage is about 12mg/m²,about 20 mg/m²,about 25 mg/m² or about 30 mg/m². In otherembodiments the dosage is 10 mg/m², 11 mg/m², 12 mg/m², 13 mg/m², 14mg/m², 15 mg/m², 16 mg/m², 17 mg/m², 18 mg/m², 19 mg/m², 20 mg/m², 21mg/m², 22 mg/m², 23 mg/m², 24 mg/m², 25 mg/m², 26 mg/m², 27 mg/m², 28mg/m², 29 mg/m², or 30 mg/m². In these embodiments the dosage amountsare administered intravenously once every four weeks i.e. 28-day cycle.

Alternatively, the dosage amount is between about 10 mg/m² to 33 mg/m².For example, the dosage is between about 11.5 mg/m² to about 12.5 mg/m².Alternatively, the dosage is between about 19.5 mg/m² to about 20.5mg/m², between about 24.5 mg/m² to about 25.5 mg/m², between about 29.5mg/m² to about 31.5 mg/m², or between about 32.5 mg/m² to about 33.5mg/m². In some embodiments, the dosage is about 12 mg/m², about 20mg/m², about 25 mg/m², about 30 mg/m² or about 33 mg/m². In otherembodiments the dosage is 10 mg/m², 11 mg/m², 12 mg/m², 13 mg/m², 14mg/m², 15 mg/m², 16 mg/m², 17 mg/m², 18 mg/m², 19 mg/m², 20 mg/m², 21mg/m², 22 mg/m², 23 mg/m², 24 mg/m², 25 mg/m², 26 mg/m², 27 mg/m², 28mg/m², 29 mg/m², 30 mg/m², 31 mg/m², 32 mg/m², or 33 mg/m². In theseembodiments the dosage amounts are administered intravenously once everyfour weeks i.e. 28-day cycle.

Alternatively, the dosage amount is between about 10 mg/m² to 36 mg/m².For example, the dosage is between about 11.5 mg/m² to about 12.5 mg/m².Alternatively, the dosage is between about 19.5 mg/m² to about 20.5mg/m², between about 24.5 mg/m² to about 25.5 mg/m², between about 29.5mg/m² to about 31.5 mg/m², between about 32.5 mg/m²to about 33.5 mg/m2,or between about 35.5 mg/m² to about 36.5 mg/m². In some embodiments,the dosage is about 12 mg/m², about 20 mg/m², about 25 mg/m², about 30mg/m², about 33 mg/m² or about 36 mg/m². In other embodiments the dosageis 10 mg/m², 11 mg/m², 12 mg/m², 13 mg/m², 14 mg/m², 15 mg/m², 16 mg/m²,17 mg/m², 18 mg/m², 19 mg/m², 20 mg/m², 21 mg/m², 22 mg/m², 23 mg/m², 24mg/m², 25 mg/m², 26 mg/m², 27 mg/m², 28 mg/m², 29 mg/m², 30 mg/m², 31mg/m², 32 mg/m², 33 mg/m², 34 mg/m², 35 mg/m², or 36 mg/m². In theseembodiments the dosage amounts are administered intravenously once everyfour weeks i.e. 28-day cycle.

Alternatively the dosage amount is between about 10 mg/m² to 40 mg/m².For example, the dosage is between about 11.5 mg/m² to about 12.5 mg/m².Alternatively the dosage is between about 19.5 mg/m² to about 20.5mg/m², between about 24.5 mg/m² to about 25.5 mg/m², between about 29.5mg/m² to about 31.5 mg/m², between about 32.5 mg/m² to about 33.5 mg/m²,between about 35.5 mg/m² to about 36.5 mg/m², between about 39.5 mg/m²toabout 41.5 mg/m². In some embodiments, the dosage is about 12 mg/m²,about 20 mg/m², about 25 mg/m², about 30 mg/m², about 33 mg/m², about 36mg/m² or about 40 mg/m². In other embodiments the dosage is 10 mg/m², 11mg/m², 12 mg/m², 13 mg/m², 14 mg/m², 15 mg/m², 16 mg/m², 17 mg/m², 18mg/m², 19 mg/m², 20 mg/m², 21 mg/m², 22 mg/m², 23 mg/m², 24 mg/m², 25mg/m², 26 mg/m², 27 mg/m², 28 mg/m², 29 mg/m², 30 mg/m², 31 mg/m², 32mg/m², 33 mg/m², 34 mg/m², 35 mg/m², 36 mg/m², 37 mg/m², 38 mg/m², 39mg/m², 40 mg/m². In these embodiments the dosage amounts areadministered intravenously once every four weeks i.e. 28-day cycle.

Alternatively the dosage amount is between about 10 mg/m² to 45 mg/m².For example, the dosage is between about 11.5 mg/m² to about 12.5 mg/m².Alternatively the dosage is between about 19.5 mg/m² to about 20.5mg/m², between about 24.5 mg/m² to about 25.5 mg/m², between about 29.5mg/m² to about 31.5 mg/m², between about 32.5 mg/m² to about 33.5 mg/m²,between about 35.5 mg/m² to about 36.5 mg/m², between about 39.5 mg/m²toabout 41.5 mg/m², between about 42.5 mg/m² to about 43.5 mg/m², betweenabout 43 mg/m² to about 45 mg/m². In some embodiments, the dosage isabout 12 mg/m², about 20 mg/m²,about 25 mg/m², about 30 mg/m², about 33mg/m², about 36 mg/m², about 40 mg/m², about 43 mg/m², or about 45mg/m². In other embodiments the dosage is 10 mg/m², 11 mg/m², 12 mg/m²,13 mg/m², 14 mg/m², 15 mg/m², 16 mg/m², 17 mg/m², 18 mg/m², 19 mg/m², 20mg/m², 21 mg/m², 22 mg/m², 23 mg/m², 24 mg/m², 25 mg/m², 26 mg/m², 27mg/m², 28 mg/m², 29 mg/m², 30 mg/m², 31 mg/m², 32 mg/m², 33 mg/m², 34mg/m², 35 mg/m², 36 mg/m², 37 mg/m², 38 mg/m², 39 mg/m², 40 mg/m², 41mg/m², 42 mg/m², 43 mg/m², 44 mg/m² or 45 mg/m². In these embodimentsthe dosage amounts are administered intravenously once every four weeksi.e. 28-day cycle.

The frequency and timing of administration, and the dosage amounts, canbe administered periodically over a cycle of administration to maintaina continuous and/or long term effect of the active agents for a desiredlength of time. The provided compositions of a NaPi2b-targeted polymerantibody-drug conjugate can be administered hourly, daily, weekly,monthly, yearly or once. The length of time of the cycle ofadministration can be empirically determined, and is dependent on thedisease to be treated, the severity of the disease, the particularpatient, and other considerations within the level of skill of thetreating physician. The length of time of treatment with a combinationtherapy provided herein can be one week, two weeks, one months, severalmonths, one year, several years or more.

For example, the frequency of administration of the NaPi2b-targetedpolymer antibody-drug conjugate is once a day, every other day, twiceweekly, once weekly, once every 2 weeks, once every 3 weeks or onceevery 4 weeks. The dosage can be divided into a plurality of cycles ofadministration during the course of treatment. For example, theNaPi2b-targeted polymer antibody-drug conjugate can be administered atthe frequency over a period of about a month, 2 months, 3 months, 4months, 5 months, 6 months, a year or more. The frequency ofadministration can be the same throughout the period of the cycle or candiffer. For example, an exemplary dosage frequency is two times a weekat least for a first week of a cycle of administration. After the firstweek, the frequency can continue at twice a week, can increase to morethan twice a week, or can be reduced to no more than once a week. It iswithin the level of a skilled person to determine the particular dosagefrequency and cycle of administration based on the particular dosagebeing administered, the disease or condition being treated, the severityof the disease or condition, the age of the subject and other similarfactors.

If disease symptoms persist in the absence of discontinued treatment,treatment can be continued for an additional length of time. Over thecourse of treatment, evidence of disease and/or treatment-relatedtoxicity or side effects can be monitored.

The cycle of administration of the NaPi2b-targeted polymer antibody-drugconjugate can be tailored to add periods of discontinued treatment inorder to provide a rest period from exposure to the agents. The lengthof time for the discontinuation of treatment can be for a predeterminedtime or can be empirically determined depending on how the patient isresponding or depending on observed side effects. For example, thetreatment can be discontinued for one week, two weeks, three weeks, onemonth or several months. Generally, the period of discontinued treatmentis built into a cycle of dosing regimen for a patient.

An exemplary dosing regimen is a treatment cycle or cycle ofadministration of 21 days or 28 days. Preferably, the dosing regimen isa treatment cycle or cycle of administration is 28 days. TheNaPi2b-targeted polymer antibody-drug conjugate disclosed herein, isadministered on day 1, followed by 20 days without dosing or isadministered on day 1, followed by 27 days without dosing. It is withinthe level of one of skill in the art to determine the precise cycle ofadministration and dosing schedule.

As noted above, the cycle of administration can be for any desiredlength of time. Hence, the 21-day cycle or 28-day cycle ofadministration can be repeated for any length of time. For example, the21-day cycle or 28-day cycle of administration can be repeated for 2months, 3, months, 4 months, 5, months, 6 months, 7 months, 8 months, 9months, 10 months, 11 months, 12 months, 1.5 years, 2 years, 2.5 years,3 years or more. It is within the level of skill of the treatingphysician to adopt a cycle of administration and dosing regimen thatmeets the needs of the patient depending on personal considerationsspecific to the patient and disease to be treated.

Measurement of NaPi2b Expression

In various aspect the invention provides a method for identifying acancer patient amenable to NaPi2b targeted therapy or monitoring thetreatment regimen by measuring the status of NaPi2b expression in atumor sample obtained from the patient.

In some embodiments, the NaPi2b diagnostic tests can be used toidentification subjects for treatment with the NaPi2b targeted polymerdrug conjugate.

The sample is derived from the subject having a cancer. The sample ofcancer cells is dissected from tissue removed or obtained from thesubject. In some embodiments, the sample is a fresh, frozen or anarchival biopsy sample.

In some embodiments, the test cell population is derived from fresh,unfrozen tissue from a biopsy sample. In other embodiments, the testcell population is derived from a primary or metastatic site. In someembodiments, the test cell population is derived from a fresh or frozentissue from a biopsy or surgical sample or ascetic fluid or pleuralfluid. In some embodiments, the test cell population is derived from afixed tissue (e.g., formalin fixation or formalin-fixedparaffin-embedded (FFPE)) from a biopsy or surgical sample or cell blockderived from a fluid specimen. The tissue sample may be frozen or fresh.

The requisite level of NaPi2b expression may be that which is identifiedby the any methods known in the art and more specifically by the methodsdescribed herein. For example, the level of NaPi2b expression can bemeasured by conducting a known immunological assay, such as an enzymeimmunoassay, radioimmunoassay, competitive immunoassay, double antibodysandwich assay, fluoroimmuno assay, ELISA, Western blotting technique,agglutination assay, cytofluorometry (e.g. flow cytometry), Fluorescencein situ hybridization (FISH), colorimetric or immunohistochemicalstaining assay (IHC) for protein expression using an antibody thatspecifically recognizes NaPi2b. Cell-based assays, such as, for example,flow cytometry (FC), immuno-histochemistry (IHC), RNA expressionanalysis or immunofluorescence (IF) are particularly desirable indetermining NaPi2b expression status, since such assay formats areclinically-suitable.

Flow cytometry (FC) may be employed to determine cell surface expressionof NaPi2b in a tumor sample before, during, and after treatment with adrug. For example, tumor cells may be analyzed by flow cytometry forNaPi2b expression, as well as for markers identifying cancer cell types,etc., if so desired. Flow cytometry may be carried out according tostandard methods. See, e.g. Chow et al., Cytometry (Communications inClinical Cytometry) 46: 72-78 (2001). Briefly and by way of example, thefollowing protocol for cytometric analysis may be employed: fixation ofthe cells with 2% paraformaldehyde for 10 minutes at 37° C. followed bypermeabilization in 90% methanol for 30 minutes on ice. Cells may thenbe stained with NaPi2b-specific antibody, washed and labeled with afluorescent-labeled secondary antibody. The cells would then be analyzedon a flow cytometer (e.g. a Beckman Coulter FC500) according to thespecific protocols of the instrument used. Such an analysis wouldidentify the level of expressed NaPi2b in the tumor.

Immunohistochemical (IHC) staining may be also employed to determine theexpression of NaPi2b in a tumor sample before, during, and aftertreatment with a drug. IHC may be carried out according to well-knowntechniques. See, e.g., ANTIBODIES; A LABORATORY MANUAL, Chapter 10,Harlow & Lane Eds., Cold Spring Harbor Laboratory (1988). Briefly, andby way of example, paraffin-embedded tissue (e.g. tumor tissue from abiopsy) is prepared for immunohistochemical staining by deparaffinizingtissue sections with xylene followed by ethanol; hydrating in water thenPBS; unmasking antigen by heating slide in sodium citrate buffer;incubating sections in hydrogen peroxide; blocking in blocking solution;incubating slide in primary polypeptide antibody and secondary antibody;and finally detecting using ABC avidin/biotin method according tomanufacturer's instructions.

Immunofluorescence (IF) assays may be also employed to determine theexpression of NaPi2b tumor sample before, during, and after treatmentwith a drug. IF may be carried out according to well-known techniques.See, e.g., J. M. Polak and S. Van Noorden (1997) INTRODUCTION TOIMMUNOCYTOCHEMISTRY, 2nd Ed.; ROYAL MICROSCOPY SOCIETY MICROSCOPYHANDBOOK 37, BioScientific/Springer-Verlag. Briefly, and by way ofexample, patient samples may be fixed in paraformaldehyde followed bymethanol, blocked with a blocking solution such as horse serum,incubated with the primary antibody against polypeptide followed by asecondary antibody labeled with a fluorescent dye such as Alexa 488 andanalyzed with an epifluorescent microscope.

Antibodies employed in the above-described assays may be advantageouslyconjugated to fluorescent dyes (e.g. Alexa488, PE), or other labels,such as quantum dots, for use in multi-parametric analyses along withother signal transduction (phospho-AKT, phospho-Erk 1/2) and/or cellmarker (cytokeratin) antibodies.

In a preferred embodiment the expression of NaPi2b in a sample from atumor is determined immunohistochemically. In another embodiment, theexpression of NaPi2b in a sample from a tumor is determinedimmunohistochemically (IHC) using the method described in U.S. Ser. No.16/136,706, which is incorporated herein in its entirety by reference.

Alternatively, the assay may include preparing RNA from the sample,optionally for use in PCR (polymerase chain reaction) or otheranalytical methodology. The PCR methodology is optionally, for example,RT-PCR (reverse transcription-PCR) or quantitative PCR, such as, forexample, real-time RT-PCR, RNA seq and the like. Alternatively, theassaying may be conducted by use of an array, such as a microarray asknown in the relevant field, such as, for example, nanostringtechnologies.

Patients are identified as being responsive to treatment, wherein thetreatment is monitored or cancer is detected by detecting and/ormeasuring the expression level of NaPi2b in a sample.

The detection/measurement of the expression level of NaPi2b isdetermined by calculating a NaPi2b score. The NaPi2b score isquantitative or semi quantitative. For example detection is scoredpathologically to arrive at a pathology score. It is contemplated thatany scoring methods known in the art may be used in the methods of theinvention. In particular, any histological scoring methods known in theart.

The methods for assessing the measurement results obtained byimmunohistochemical staining assays include, for example, the H-scoremethod. The H-score is determined by the following calculation formula(Am J Clin Pathol. 1988; 90 (3): 233-9). H-Score=((% at <1+)X 0) +((% at1+)X 1)+((% at 2+)X 2)+((% at 3+) X3) where staining intensity 0 isunstained; staining intensity 1 is weak staining; staining intensity 2is moderate staining; and staining intensity 3 is strong staining.

In assessment by the H-score method, only cancer cell portions are used.For negative or positive controls for staining intensity, formalin-fixedparaffin-embedded cell lines or xenografts (lines whose proteinexpression levels are known in advance) may be employed. When there areno control specimens, a plurality of specimens are assessedsimultaneously to confirm the overall distribution of staining intensityof the specimens, and then staining intensity may be set.

In addition to the H-score method, other scoring methodsknown in theart, such as, for example, the Allred method (Harvey, et al. Journal ofClinical Oncology 17, No. 5 (May 1999) 1474-1474), can also be used.Cut-off points are required to be set in each method. Allred score=scoreof percentage of positive cells+staining intensity score.

The disclosure also provides kits and/or methods for identifying orotherwise refining, e.g., stratifying, a patient population suitable fortherapeutic administration of a NaPi2b-targeted antibody-drug conjugatesdisclosed herein by identifying the NaPi2b score of the subject prior totreatment with a NaPi2b-targeted antibody-drug conjugate disclosedherein. In some embodiments, the test cell population is derived fromfresh, unfrozen tissue from a biopsy sample. In some embodiments, thetest cell population is derived from a primary or metastatic site. Insome embodiments, the test cell population is derived from a frozentissue from a biopsy or surgical sample or ascetic fluid or pleuralfluid. In some embodiments, the test cell population is derived from afixed tissue (e.g., formalin fixation) from a biopsy or surgical sample.The IHC test measures the amount of NaPi2b receptor protein on thesurface of cells in a cancer tissue sample

DEFINITIONS

Unless otherwise defined, scientific and technical terms used inconnection with the present disclosure shall have the meanings that arecommonly understood by those of ordinary skill in the art. Further,unless otherwise required by context, singular terms shall includepluralities and plural terms shall include the singular. Generally,nomenclatures utilized in connection with, and techniques of, cell andtissue culture, molecular biology, and protein and oligo- orpolynucleotide chemistry and hybridization described herein are thosewell-known and commonly used in the art. Standard techniques are usedfor recombinant DNA, oligonucleotide synthesis, and tissue culture andtransformation (e.g., electroporation, lipofection). Enzymatic reactionsand purification techniques are performed according to manufacturer'sspecifications or as commonly accomplished in the art or as describedherein. The foregoing techniques and procedures are generally performedaccording to conventional methods well known in the art and as describedin various general and more specific references that are cited anddiscussed throughout the present specification. See e.g., Sambrook etal. Molecular Cloning: A Laboratory Manual (2d ed., Cold Spring HarborLaboratory Press, Cold Spring Harbor, N.Y. (1989)). The nomenclaturesutilized in connection with, and the laboratory procedures andtechniques of, analytical chemistry, synthetic organic chemistry, andmedicinal and pharmaceutical chemistry described herein are thosewell-known and commonly used in the art. Standard techniques are usedfor chemical syntheses, chemical analyses, pharmaceutical preparation,formulation, and delivery, and treatment of patients.

As utilized in accordance with the present disclosure, the followingterms, unless otherwise indicated, shall be understood to have thefollowing meanings:

As used herein, the terms “NaPi2b” (also known as sodium-dependentphosphate transport protein 2B, SLC34A2, NaPiIIb, Npt2, Na(+)-dependentphosphate cotransporter 2B; sodium/phosphate cotransporter 2B; Na(+)/Picotransporter 2B; NaPi3b; solute carrier family 34 member 2), when usedherein, refers to human NaPi2b (e.g., GenBank Accession No. O95436.3)and includes any variants, isoforms and species homologs of NaPi2b whichare naturally expressed by cells, including tumor cells, or areexpressed on cells transfected with the NaPi2b gene. These terms aresynonymous and may be used interchangeably.

As used herein, the term “NaPi2b antibody” or “anti-NaPi2b antibody” isan antibody which binds specifically to the extracellular region ofSLC34A2.

When used herein in the context of two or more antibodies, the term“competes with” or “cross-competes with” indicates that the two or moreantibodies compete for binding to NaPi2b, e.g., compete for NaPi2bbinding in any art-recognized assay. An antibody “blocks” or“cross-blocks” one or more other antibodies from binding to NaPi2b ifthe antibody competes with the one or more other antibodies 25% or more,with 25%-74% representing “partial block” and 75%-100% representing“full block”, as determined using any art-recognized assay. For somepairs of antibodies, competition or blocking in any art-recognized assayis only observed when one antibody is coated on the plate and the otheris used to compete, and not vice versa. Unless otherwise defined ornegated by context, the terms “competes with”, “cross-competes with”,“blocks” or “cross-blocks” when used herein is also intended to coversuch pairs of antibodies

As used herein, the term “antibody” refers to immunoglobulin moleculesand immunologically active portions of immunoglobulin (Ig) molecules,i.e., molecules that contain an antigen binding site that specificallybinds (immunoreacts with) an antigen. By “specifically bind” or“immunoreacts with” “or directed against” is meant that the antibodyreacts with one or more antigenic determinants of the desired antigenand does not react with other polypeptides or binds at much loweraffinity (K_(d)>10⁻⁶). Antibodies include, but are not limited to,polyclonal, monoclonal and chimeric antibodies.

The basic antibody structural unit is known to comprise a tetramer. Eachtetramer is composed of two identical pairs of polypeptide chains, eachpair having one “light” (about 25 kDa) and one “heavy” chain (about50-70 kDa). The amino-terminal portion of each chain includes a variableregion of about 100 to 110 or more amino acids primarily responsible forantigen recognition. The carboxy-terminal portion of each chain definesa constant region primarily responsible for effector function. Ingeneral, antibody molecules obtained from humans relate to any of theclasses IgG, IgM, IgA, IgE and IgD, which differ from one another by thenature of the heavy chain present in the molecule. Certain classes havesubclasses as well, such as IgG₁, IgG₂, and others. Furthermore, inhumans, the light chain may be a kappa chain or a lambda chain.

The term “monoclonal antibody” (mAb) or “monoclonal antibodycomposition”, as used herein, refers to a population of antibodymolecules that contain only one molecular species of antibody moleculeconsisting of a unique light chain gene product and a unique heavy chaingene product. In particular, the complementarity determining regions(CDRs) of the monoclonal antibody are identical in all the molecules ofthe population. mAbs contain an antigen binding site capable ofimmunoreacting with a particular epitope of the antigen characterized bya unique binding affinity for it.

In general, antibody molecules obtained from humans relate to any of theclasses IgG, IgM, IgA, IgE and IgD, which differ from one another by thenature of the heavy chain present in the molecule. Certain classes havesubclasses as well, such as IgG₁, IgG₂, and others. Furthermore, inhumans, the light chain may be a kappa chain or a lambda chain.

The term “antigen-binding site” or “binding portion” refers to the partof the immunoglobulin molecule that participates in antigen binding. Theantigen binding site is formed by amino acid residues of the N-terminalvariable (“V”) regions of the heavy (“H”) and light (“L”) chains. Threehighly divergent stretches within the V regions of the heavy and lightchains, referred to as “hypervariable regions,” are interposed betweenmore conserved flanking stretches known as “framework regions,” or“FRs”. Thus, the term “FR” refers to amino acid sequences which arenaturally found between, and adjacent to, hypervariable regions inimmunoglobulins. In an antibody molecule, the three hypervariableregions of a light chain and the three hypervariable regions of a heavychain are disposed relative to each other in three dimensional space toform an antigen-binding surface. The antigen-binding surface iscomplementary to the three-dimensional surface of a bound antigen, andthe three hypervariable regions of each of the heavy and light chainsare referred to as “complementarity-determining regions,” or “CDRs.” Theassignment of amino acids to each domain is in accordance with thedefinitions of Kabat Sequences of Proteins of Immunological Interest(National Institutes of Health, Bethesda, Md. (1987 and 1991)), orChothia & Lesk J. Mol. Biol. 196:901-917 (1987), Chothia et al. Nature342:878-883 (1989).

As used herein, the term “epitope” includes any protein determinantcapable of specific binding to an immunoglobulin or fragment thereof, ora T-cell receptor. The term “epitope” includes any protein determinantcapable of specific binding to an immunoglobulin or T-cell receptor.Epitopic determinants usually consist of chemically active surfacegroupings of molecules such as amino acids or sugar side chains andusually have specific three dimensional structural characteristics, aswell as specific charge characteristics. An antibody is said tospecifically bind an antigen when the dissociation constant is ≤1 μM;e.g., ≤100 nM, preferably ≤10 nM and more preferably ≤1 nM.

The term “polypeptide” is used herein as a generic term to refer tonative protein, fragments, or analogs of a polypeptide sequence. Hence,native protein fragments, and analogs are species of the polypeptidegenus. The term “naturally-occurring” as used herein as applied to anobject refers to the fact that an object can be found in nature. Forexample, a polypeptide or polynucleotide sequence that is present in anorganism (including viruses) that can be isolated from a source innature and which has not been intentionally modified by man in thelaboratory or otherwise is naturally-occurring.

The following terms are used to describe the sequence relationshipsbetween two or more polynucleotide or amino acid sequences: “referencesequence”, “comparison window”, “sequence identity”, “percentage ofsequence identity”, and “substantial identity”. A “reference sequence”is a defined sequence used as a basis for a sequence comparison areference sequence may be a subset of a larger sequence, for example, asa segment of a full-length cDNA or gene sequence given in a sequencelisting or may comprise a complete cDNA or gene sequence. Generally, areference sequence is at least 18 nucleotides or 6 amino acids inlength, frequently at least 24 nucleotides or 8 amino acids in length,and often at least 48 nucleotides or 16 amino acids in length. Since twopolynucleotides or amino acid sequences may each (1) comprise a sequence(i.e., a portion of the complete polynucleotide or amino acid sequence)that is similar between the two molecules, and (2) may further comprisea sequence that is divergent between the two polynucleotides or aminoacid sequences, sequence comparisons between two (or more) molecules aretypically performed by comparing sequences of the two molecules over a“comparison window” to identify and compare local regions of sequencesimilarity. A “comparison window”, as used herein, refers to aconceptual segment of at least 18 contiguous nucleotide positions or 6amino acids wherein a polynucleotide sequence or amino acid sequence maybe compared to a reference sequence of at least 18 contiguousnucleotides or 6 amino acid sequences and wherein the portion of thepolynucleotide sequence in the comparison window may comprise additions,deletions, substitutions, and the like (i.e., gaps) of 20 percent orless as compared to the reference sequence (which does not compriseadditions or deletions) for optimal alignment of the two sequences.Optimal alignment of sequences for aligning a comparison window may beconducted by the local homology algorithm of Smith and Waterman Adv.Appl. Math. 2:482 (1981), by the homology alignment algorithm ofNeedleman and Wunsch J. Mol. Biol. 48:443 (1970), by the search forsimilarity method of Pearson and Lipman Proc. Natl. Acad. Sci. (U.S.A.)85:2444 (1988), by computerized implementations of these algorithms(GAP, BESTFIT, FASTA, and TFASTA in the Wisconsin Genetics SoftwarePackage Release 7.0, (Genetics Computer Group, 575 Science Dr., Madison,Wis.), Geneworks, or MacVector software packages), or by inspection, andthe best alignment (i.e., resulting in the highest percentage ofhomology over the comparison window) generated by the various methods isselected.

The term “sequence identity” means that two polynucleotide or amino acidsequences are identical (i.e., on a nucleotide-by-nucleotide orresidue-by-residue basis) over the comparison window. The term“percentage of sequence identity” is calculated by comparing twooptimally aligned sequences over the window of comparison, determiningthe number of positions at which the identical nucleic acid base (e.g.,A, T, C, G, U or I) or residue occurs in both sequences to yield thenumber of matched positions, dividing the number of matched positions bythe total number of positions in the comparison window (i.e., the windowsize), and multiplying the result by 100 to yield the percentage ofsequence identity. The terms “substantial identity” as used hereindenotes a characteristic of a polynucleotide or amino acid sequence,wherein the polynucleotide or amino acid comprises a sequence that hasat least 85 percent sequence identity, preferably at least 90 to 95percent sequence identity, more usually at least 99 percent sequenceidentity as compared to a reference sequence over a comparison window ofat least 18 nucleotide (6 amino acid) positions, frequently over awindow of at least 24-48 nucleotide (8-16 amino acid) positions, whereinthe percentage of sequence identity is calculated by comparing thereference sequence to the sequence which may include deletions oradditions which total 20 percent or less of the reference sequence overthe comparison window. The reference sequence may be a subset of alarger sequence.

As used herein, the twenty conventional amino acids and theirabbreviations follow conventional usage. See Immunology—A Synthesis (2ndEdition, E. S. Golub and D. R. Green, Eds., Sinauer Associates,Sunderland7 Mass. (1991)). Stereoisomers (e.g., D-amino acids) of thetwenty conventional amino acids, unnatural amino acids such as α-,α-disubstituted amino acids, N-alkyl amino acids, lactic acid, and otherunconventional amino acids may also be suitable components forpolypeptides of the present disclosure. Examples of unconventional aminoacids include: 4 hydroxyproline, γ-carboxyglutamate,ε-N,N,N-trimethyllysine, ε-N-acetyllysine, O-phosphoserine,N-acetylserine, N-formylmethionine, 3-methylhistidine, 5-hydroxylysine,σ-N-methylarginine, and other similar amino acids and imino acids (e.g.,4-hydroxyproline). In the polypeptide notation used herein, theleft-hand direction is the amino terminal direction and the right-handdirection is the carboxy-terminal direction, in accordance with standardusage and convention.

Similarly, unless specified otherwise, the left-hand end ofsingle-stranded polynucleotide sequences is the 5′ end the left-handdirection of double-stranded polynucleotide sequences is referred to asthe 5′ direction. The direction of 5′ to 3′ addition of nascent RNAtranscripts is referred to as the transcription direction sequenceregions on the DNA strand having the same sequence as the RNA and whichare 5′ to the 5′ end of the RNA transcript are referred to as “upstreamsequences”, sequence regions on the DNA strand having the same sequenceas the RNA and which are 3′ to the 3′ end of the RNA transcript arereferred to as “downstream sequences”.

As applied to polypeptides, the term “substantial identity” means thattwo peptide sequences, when optimally aligned, such as by the programsGAP or BESTFIT using default gap weights, share at least 80 percentsequence identity, preferably at least 90 percent sequence identity,more preferably at least 95 percent sequence identity, and mostpreferably at least 99 percent sequence identity.

Preferably, residue positions which are not identical differ byconservative amino acid substitutions.

Conservative amino acid substitutions refer to the interchangeability ofresidues having similar side chains. For example, a group of amino acidshaving aliphatic side chains is glycine, alanine, valine, leucine, andisoleucine; a group of amino acids having aliphatic-hydroxyl side chainsis serine and threonine; a group of amino acids having amide-containingside chains is asparagine and glutamine; a group of amino acids havingaromatic side chains is phenylalanine, tyrosine, and tryptophan; a groupof amino acids having basic side chains is lysine, arginine, andhistidine; and a group of amino acids having sulfur-containing sidechains is cysteine and methionine. Preferred conservative amino acidssubstitution groups are valine-leucine-isoleucine,phenylalanine-tyrosine, lysine-arginine, alanine valine,glutamic-aspartic, and asparagine-glutamine.

As discussed herein, minor variations in the amino acid sequences ofantibodies or immunoglobulin molecules are contemplated as beingencompassed by the present disclosure, providing that the variations inthe amino acid sequence maintain at least 75%, more preferably at least80%, 90%, 95%, and most preferably 99%. In particular, conservativeamino acid replacements are contemplated. Conservative replacements arethose that take place within a family of amino acids that are related intheir side chains. Genetically encoded amino acids are generally dividedinto families: (1) acidic amino acids are aspartate, glutamate; (2)basic amino acids are lysine, arginine, histidine; (3) non-polar aminoacids are alanine, valine, leucine, isoleucine, proline, phenylalanine,methionine, tryptophan, and (4) uncharged polar amino acids are glycine,asparagine, glutamine, cysteine, serine, threonine, tyrosine. Thehydrophilic amino acids include arginine, asparagine, aspartate,glutamine, glutamate, histidine, lysine, serine, and threonine. Thehydrophobic amino acids include alanine, cysteine, isoleucine, leucine,methionine, phenylalanine, proline, tryptophan, tyrosine and valine.Other families of amino acids include (i) serine and threonine, whichare the aliphatic-hydroxy family; (ii) asparagine and glutamine, whichare the amide containing family; (iii) alanine, valine, leucine andisoleucine, which are the aliphatic family; and (iv) phenylalanine,tryptophan, and tyrosine, which are the aromatic family. For example, itis reasonable to expect that an isolated replacement of a leucine withan isoleucine or valine, an aspartate with a glutamate, a threonine witha serine, or a similar replacement of an amino acid with a structurallyrelated amino acid will not have a major effect on the binding orproperties of the resulting molecule, especially if the replacement doesnot involve an amino acid within a framework site. Whether an amino acidchange results in a functional peptide can readily be determined byassaying the specific activity of the polypeptide derivative. Assays aredescribed in detail herein. Fragments or analogs of antibodies orimmunoglobulin molecules can be readily prepared by those of ordinaryskill in the art. Preferred amino- and carboxy-termini of fragments oranalogs occur near boundaries of functional domains. Structural andfunctional domains can be identified by comparison of the nucleotideand/or amino acid sequence data to public or proprietary sequencedatabases. Preferably, computerized comparison methods are used toidentify sequence motifs or predicted protein conformation domains thatoccur in other proteins of known structure and/or function. Methods toidentify protein sequences that fold into a known three-dimensionalstructure are known. Bowie et al. Science 253:164 (1991). Thus, theforegoing examples demonstrate that those of skill in the art canrecognize sequence motifs and structural conformations that may be usedto define structural and functional domains in accordance with thedisclosure.

Preferred amino acid substitutions are those which: (1) reducesusceptibility to proteolysis, (2) reduce susceptibility to oxidation,(3) alter binding affinity for forming protein complexes, (4) alterbinding affinities, and (4) confer or modify other physicochemical orfunctional properties of such analogs. Analogs can include variousmuteins of a sequence other than the naturally-occurring peptidesequence. For example, single or multiple amino acid substitutions(preferably conservative amino acid substitutions) may be made in thenaturally-occurring sequence (preferably in the portion of thepolypeptide outside the domain(s) forming intermolecular contacts. Aconservative amino acid substitution should not substantially change thestructural characteristics of the parent sequence (e.g., a replacementamino acid should not tend to break a helix that occurs in the parentsequence, or disrupt other types of secondary structure thatcharacterizes the parent sequence). Examples of art-recognizedpolypeptide secondary and tertiary structures are described in Proteins,Structures and Molecular Principles (Creighton, Ed., W. H. Freeman andCompany, New York (1984)); Introduction to Protein Structure (C. Brandenand J. Tooze, eds., Garland Publishing, New York, N.Y. (1991)); andThornton et at. Nature 354:105 (1991).

Peptide analogs are commonly used in the pharmaceutical industry asnon-peptide drugs with properties analogous to those of the templatepeptide. These types of non-peptide compound are termed “peptidemimetics” or “peptidomimetics”. Fauchere, J. Adv. Drug Res. 15:29(1986), Veber and Freidinger TINS p.392 (1985); and Evans et al. J. Med.Chem. 30:1229 (1987). Such compounds are often developed with the aid ofcomputerized molecular modeling. Peptide mimetics that are structurallysimilar to therapeutically useful peptides may be used to produce anequivalent therapeutic or prophylactic effect. Generally,peptidomimetics are structurally similar to a paradigm polypeptide(i.e., a polypeptide that has a biochemical property or pharmacologicalactivity), such as human antibody, but have one or more peptide linkagesoptionally replaced by a linkage selected from the group consisting of:—CH₂NH—, —CH₂S—, —CH₂—CH₂—, —CH═CH-(cis and trans), —COCH₂—, CH(OH)CH₂—,and —CH₂SO—, by methods well known in the art. Systematic substitutionof one or more amino acids of a consensus sequence with a D-amino acidof the same type (e.g., D-lysine in place of L-lysine) may be used togenerate more stable peptides. In addition, constrained peptidescomprising a consensus sequence or a substantially identical consensussequence variation may be generated by methods known in the art (Rizoand Gierasch Ann. Rev. Biochem. 61:387 (1992)); for example, by addinginternal cysteine residues capable of forming intramolecular disulfidebridges which cyclize the peptide.

The term “agent” is used herein to denote a chemical compound, a mixtureof chemical compounds, a biological macromolecule, or an extract madefrom biological materials.

As used herein, the terms “label” or “labeled” refers to incorporationof a detectable marker, e.g., by incorporation of a radiolabeled aminoacid or attachment to a polypeptide of biotinyl moieties that can bedetected by marked avidin (e.g., streptavidin containing a fluorescentmarker or enzymatic activity that can be detected by optical orcalorimetric methods). In certain situations, the label or marker canalso be therapeutic. Various methods of labeling polypeptides andglycoproteins are known in the art and may be used. Examples of labelsfor polypeptides include, but are not limited to, the following:radioisotopes or radionuclides (e.g., ³H, ¹⁴C, ¹⁵N, ³⁵S, ⁹⁰Y, ⁹⁹Tc,¹¹¹In, ¹²⁵I, ¹³¹I), fluorescent labels (e.g., FITC, rhodamine,lanthanide phosphors), enzymatic labels (e.g., horseradish peroxidase,p-galactosidase, luciferase, alkaline phosphatase), chemiluminescent,biotinyl groups, predetermined polypeptide epitopes recognized by asecondary reporter (e.g., leucine zipper pair sequences, binding sitesfor secondary antibodies, metal binding domains, epitope tags). In someembodiments, labels are attached by spacer arms of various lengths toreduce potential steric hindrance. The term “pharmaceutical agent ordrug” as used herein refers to a chemical compound or compositioncapable of inducing a desired therapeutic effect when properlyadministered to a patient.

Other chemistry terms herein are used according to conventional usage inthe art, as exemplified by The McGraw-Hill Dictionary of Chemical Terms(Parker, S., Ed., McGraw-Hill, San Francisco (1985)).

As used herein, “substantially pure” means an object species is thepredominant species present (i.e., on a molar basis it is more abundantthan any other individual species in the composition), and preferably asubstantially purified fraction is a composition wherein the objectspecies comprises at least about 50 percent (on a molar basis) of allmacromolecular species present.

Generally, a substantially pure composition will comprise more thanabout 80 percent of all macromolecular species present in thecomposition, more preferably more than about 85%, 90%, 95%, and 99%.Most preferably, the object species is purified to essential homogeneity(contaminant species cannot be detected in the composition byconventional detection methods) wherein the composition consistsessentially of a single macromolecular species.

The use of the articles “a”, “an”, and “the” in both the followingdescription and claims are to be construed to cover both the singularand the plural, unless otherwise indicated herein or clearlycontradicted by context. The terms “comprising”, “having”, “being of” asin “being of a chemical formula”, “including”, and “containing” are tobe construed as open terms (i.e., meaning “including but not limitedto”) unless otherwise noted. For example, a polymeric scaffold of acertain formula includes all the monomer units shown in the formula andmay also include additional monomer units not shown in the formula.Additionally, whenever “comprising” or another open-ended term is usedin an embodiment, it is to be understood that the same embodiment can bemore narrowly claimed using the intermediate term “consistingessentially of” or the closed term “consisting of.”

The term “about”, “approximately”, or “approximate”, when used inconnection with a numerical value, means that a collection or range ofvalues is included. For example, “about X” includes a range of valuesthat are ±20%, ±10%, ±5%, ±2%, ±1%, ±0.5%, ±0.2%, or ±0.1% of X, where Xis a numerical value. In one embodiment, the term “about” refers to arange of values which are 5% more or less than the specified value. Inanother embodiment, the term “about” refers to a range of values whichare 2% more or less than the specified value. In another embodiment, theterm “about” refers to a range of values which are 1% more or less thanthe specified value.

Recitation of ranges of values are merely intended to serve as ashorthand method of referring individually to each separate valuefalling within the range, unless otherwise indicated herein, and eachseparate value is incorporated into the specification as if it wereindividually recited herein. A range used herein, unless otherwisespecified, includes the two limits of the range. For example, theexpressions “x being an integer between 1 and 6” and “x being an integerof 1 to 6” both mean “x being 1, 2, 3, 4, 5, or 6”, i.e., the terms“between X and Y” and “range from X to Y, are inclusive of X and Y andthe integers there between.

All methods described herein can be performed in any suitable orderunless otherwise indicated herein or otherwise clearly contradicted bycontext. The use of any and all examples, or exemplary language (e.g.,“such as”) provided herein, is intended merely to better illustrate theinvention and is not to be construed as a limitation on the scope of theclaims unless explicitly otherwise claimed. No language in thespecification is to be construed as indicating that any non-claimedelement is essential to what is claimed.

“Polymeric Carrier or scaffold”: The term polymeric carrier or scaffold,as used herein, refers to a polymer or a modified polymer, which issuitable for covalently attaching to or can be covalently attached toone or more drug molecules with a designated linker and/or one or morePBRMs with a designated linker.

“Physiological conditions”: The phrase “physiological conditions”, asused herein, relates to the range of chemical (e.g., pH, ionic strength)and biochemical (e.g., enzyme concentrations) conditions likely to beencountered in the extracellular fluids of living tissues. For mostnormal tissues, the physiological pH ranges from about 7.0 to 7.4.Circulating blood plasma and normal interstitial liquid representtypical examples of normal physiological conditions.

“Drug”: As used herein, the term “drug” refers to a compound which isbiologically active and provides a desired physiological effectfollowing administration to a subject in need thereof (e.g., an activepharmaceutical ingredient).

“Cytotoxic”: As used herein the term “cytotoxic” means toxic to cells ora selected cell population (e.g., cancer cells). The toxic effect mayresult in cell death and/or lysis. In certain instances, the toxiceffect may be a sublethal destructive effect on the cell, e.g., slowingor arresting cell growth. In order to achieve a cytotoxic effect, thedrug or prodrug may be selected from a group consisting of a DNAdamaging agent, a microtubule disrupting agent, or a cytotoxic proteinor polypeptide, amongst others.

“PHF” refers to poly(1-hydroxymethylethylene hydroxymethyl-formal).

As used herein, the terms “polymer unit”, “monomeric unit”, “monomer”,“monomer unit”, “unit” all refer to a repeatable structural unit in apolymer.

As used herein, “molecular weight” or “MW” of a polymer or polymericcarrier/scaffold or polymer conjugates refers to the weight averagemolecular weight of the unmodified polymer unless otherwise specified.

As used herein, “dosing regimen” or “dosage regimen” refers to theamount of agent, for example, the composition containing anNaPi2b-targeted polymer antibody-drug conjugate, administered, and thefrequency of administration. The dosing regimen is a function of thedisease or condition to be treated, and thus can vary.

As used herein, “frequency” of administration refers to the time betweensuccessive administrations of treatment. For example, frequency can bedays, weeks or months. For example, frequency can be more than onceweekly, for example, twice a week, three times a week, four times aweek, five times a week, six times a week or daily. Frequency also canbe one, two, three or four weeks. The particular frequency is a functionof the particular disease or condition treated. Generally, frequency ismore than once weekly, and generally is twice weekly.

As used herein, a “cycle of administration” refers to the repeatedschedule of the dosing regimen of administration of the enzyme and/or asecond agent that is repeated over successive administrations. Forexample, an exemplary cycle of administration is a 28 day cycle withadministration twice weekly for three weeks, followed by one-week ofdiscontinued dosing. A preferred cycle of administration is a 21 daycycle with administration once every 21 days (i.e., 3 weeks) or a 28 daycycle with administration once every 28 days (i.e., 4 weeks)

As used herein, when referencing dosage based on mg/kg of the subject,an average human subject is considered to have a mass of about 70 kg-75kg, such as 70 kg and a body surface area (BSA) of 1.73 m².

As used herein, amelioration of the symptoms of a particular disease ordisorder by a treatment, such as by administration of a pharmaceuticalcomposition or other therapeutic, refers to any lessening, whetherpermanent or temporary, lasting or transient, of the symptoms or,adverse effects of a condition, such as, for example, reduction ofadverse effects associated with or that occur upon administration of anNaPi2b-targeted polymer antibody-drug conjugate.

As used herein, when referencing dosage based on “body surface area”(BSA; m²) is the measured or calculated surface area of a human body.For many clinical purposes BSA is a better indicator of metabolic massthan body weight because it is less affected by abnormal adipose mass.Various calculations have been published to arrive at the BSA withoutdirect measurement. In the following formulae, BSA is in m², W is massin kg, and H is height in cm. The most widely used is the Du Bois, DuBois formula: BSA=0.007184×W^(0.425)×H^(0.725). Other methods ofdetermining BSA include for example, the Mosteller, Haycock, Gehan andGeorge, Boyd, Fujimoto, Takahira, Shuter and Aslani or Schlich formulas

As used herein, “treating” or “treat” describes the management and careof a patient for the purpose of combating a disease, condition, ordisorder and includes the administration of a conjugate of thedisclosure, or a pharmaceutical composition thereof in combination withan immunomodulatory therapy, e.g., an immuno-oncology agent such as animmune checkpoint inhibitor, to alleviate the symptoms or complicationsof a disease, condition or disorder, or to eliminate the disease,condition or disorder.

As used herein, “prevention” or “prophylaxis” refers to reduction in therisk of developing a disease or condition, or reduction or eliminationof the onset of the symptoms or complications of the disease, conditionor disorder.

The term “effective amount” or “sufficient amount”, as it refers to anactive agent, refers to the amount necessary to elicit the desiredbiological response. As used herein, a “therapeutically effectiveamount” or a “therapeutically effective dose” refers to an amount orquantity of an agent, compound, material, or composition containing acompound that is at least sufficient to produce a detectable therapeuticeffect. The effect can be detected by any assay method known in the art.The precise effective amount for a subject will depend upon thesubject's body weight, size, and health; the nature and extent of thecondition; and the therapeutic selected for administration.

A “subject” includes a mammal. The mammal can be e.g., any mammal, e.g.,a human, primate, bird, mouse, rat, fowl, dog, cat, cow, horse, goat,camel, sheep or a pig. Preferably, the mammal is a human.

As used herein, “unit dose form” or “unit dosage form” refers tophysically discrete units suitable for human and animal subjects andpackaged individually as is known in the art.

As used herein, a single dosage formulation refers to a formulation as asingle dose.

As used herein, “temporal proximity” refers to that administration ofone therapeutic agent (e.g., a NaPi2b-targeted polymer antibody-drugconjugate disclosed herein) occurs within a time period before or afterthe administration of another therapeutic agent (e.g., an immunecheckpoint inhibitor disclosed herein), such that the therapeutic effectof the one therapeutic agent overlaps with the therapeutic effect of theanother therapeutic agent. In some embodiments, the therapeutic effectof the one therapeutic agent completely overlaps with the therapeuticeffect of the another therapeutic agent. In some embodiments, “temporalproximity” means that administration of one therapeutic agent occurswithin a time period before or after the administration of anothertherapeutic agent, such that there is a synergistic effect between theone therapeutic agent and the another therapeutic agent. “Temporalproximity” may vary according to various factors, including but notlimited to, the age, gender, weight, genetic background, medicalcondition, disease history, and treatment history of the subject towhich the therapeutic agents are to be administered; the disease orcondition to be treated or ameliorated; the therapeutic outcome to beachieved; the dosage, dosing frequency, and dosing duration of thetherapeutic agents; the pharmacokinetics and pharmacodynamics of thetherapeutic agents; and the route(s) through which the therapeuticagents are administered. In some embodiments, “temporal proximity” meanswithin 15 minutes, within 30 minutes, within an hour, within two hours,within four hours, within six hours, within eight hours, within 12hours, within 18 hours, within 24 hours, within 36 hours, within 2 days,within 3 days, within 4 days, within 5 days, within 6 days, within aweek, within 2 weeks, within 3 weeks, within 4 weeks, with 6 weeks, orwithin 8 weeks. In some embodiments, multiple administration of onetherapeutic agent can occur in temporal proximity to a singleadministration of another therapeutic agent. In some embodiments,temporal proximity may change during a treatment cycle or within adosing regimen.

As used herein a “kit” refers to a combination of components, such as acombination of the compositions herein and another item for a purposeincluding, but not limited to, reconstitution, activation andinstruments/devices for delivery, administration, diagnosis andassessment of a biological activity or property. Kits optionally includeinstructions of use.

The present disclosure is intended to include all isotopes of atomsoccurring in the present compounds. Isotopes include those atoms havingthe same atomic number but different mass numbers. By way of generalexample and without limitation, isotopes of hydrogen include tritium anddeuterium. Isotopes of carbon include C-13 and C-14.

The present disclosure is intended to include all isomers of thecompound, which refers to and includes, optical isomers, and tautomericisomers, where optical isomers include enantiomers and diastereomers,chiral isomers and non-chiral isomers, and the optical isomers includeisolated optical isomers as well as mixtures of optical isomersincluding racemic and non-racemic mixtures; where an isomer may be inisolated form or in a mixture with one or more other isomers.

OTHER EMBODIMENTS

All publications and patent documents cited herein are incorporatedherein by reference as if each such publication or document wasspecifically and individually indicated to be incorporated herein byreference. Citation of publications and patent documents is not intendedas an admission that any is pertinent prior art, nor does it constituteany admission as to the contents or date of the same. The inventionhaving now been described by way of written description, those of skillin the art will recognize that the invention can be practiced in avariety of embodiments and that the foregoing description and examplesbelow are for purposes of illustration and not limitation of the claimsthat follow.

EXAMPLES

The following examples are illustrative and are not intended to belimiting and it will be readily understood by one of skill in the artthat other reagents or methods may be utilized.

ABBREVIATIONS

The following abbreviations are used in the reaction schemes andsynthetic examples, which follow. This list is not meant to be anall-inclusive list of abbreviations used in the application asadditional standard abbreviations, which are readily understood by thoseskilled in the art of organic synthesis, can also be used in thesynthetic schemes and examples.

AF-HPA Auristatin F-hydroxypropyl amide

BSA Body surface area

CR Complete response

DCR Disease control rate

DES Dose escalation

DLT Dose limiting toxicity

EXP Cohort expansion

IHC Immunohistochemistry

IV Intravenous

MTD Maximum tolerated dose

NSCLC Non-small cell lung cancer

ORR Objective response rate

PBS Phosphate buffered saline

PE Physical examination

PHF poly(1-hydroxymethylethylene hydroxylmethylformal)

PR Partial response

RP2D Recommended Phase 2 dose

SD Stable disease

SRC Safety Review Committee

SRM Safety Review Meeting

TRAE Treatment-related adverse events

GENERAL INFORMATION

XMT-1536 was prepared as described in US Application No. 2017/0266311.

AF-HPA was prepared as described in U.S. Pat. No. 8,808,679(B2)

CDRs were identified by the Kabat numbering scheme.

Example 1: Assessment of Safety and Tolerability of XMT-1536 StudyDesign

The study presented herein is an open label, multi-center Phase 1b studyof XMT-1536 administered as an intravenous infusion once every three orfour weeks. The dose escalation (DES) part of the study will establishthe maximum tolerated dose (MTD) or recommended Phase 2 dose (RP2D) forXMT-1536 in patients with a number of tumor types likely to expressNaPi2b, with a focus on patients with platinum-resistant ovarian cancerand non-squamous non-small cell lung cancer (NSCLC). The MTD is definedas the highest dose of XMT-1536 that does not cause unacceptabletoxicities defined by the protocol-specific dose limit-toxicitycriteria. The RP2D may differ from the MTD and may take into accounttolerability, pharmacokinetic and pharmacodynamic measures, and efficacyof XMT-1536 based on RECIST 1.1. After the first cycle, patients maycontinue to receive XMT-1536 until disease progression provided the drugis well-tolerated and patients continue to derive clinical benefit inthe opinion of the Investigator. The DES segment of the study utilizedan accelerated titration design. The first 3-week cycle of treatmentconstitutes the dose limiting toxicity (DLT) evaluation period. For thefirst two dose levels, a minimum of 1 patient was treated at each doselevel. If this patient experiences a treatment-emergent adverse event ofGrade 2 or higher, 2 additional patients will be added at this doselevel, and the study will subsequently follow a 3+3 design, as describedbelow. If this patient does not experience a Grade 2 or highertreatment-related event or a DLT during the DLT evaluation period, andthe Safety Review Committee (SRC) agrees this was a reasonablywell-tolerated dose, enrollment will commence at the next dose level.Starting with Dose Level 3, the study will follow a standard 3+3 design,with 3 patients enrolled initially at each dose level. If none of the 3patients experiences a DLT during the evaluation period and the SRCagrees this was a reasonably well tolerated dose, 3 patients will beenrolled at the next dose level. However, in the event of 1 DLT, 3additional patients will be enrolled at the same dose level. Any doselevel with 2 or more DLTs will be considered to exceed the MTD andsubsequent patients will be enrolled at lower dose level(s). After thefirst cycle, patients may continue to receive XMT-1536 until diseaseprogression if the drug is well-tolerated and patients continue toderive clinical benefit in the opinion of the Investigator.

After completion of DES, the EXP segment of the study will be opened inthree cohorts of patients. Cohort 1 will contain patients withplatinum-resistant ovarian cancer. Cohort 2 will contain patients withnon-squamous NSCLC. Cohort 3 will contain patients with papillarythyroid carcinoma, endometrial carcinoma, papillary renal cellcarcinoma, or salivary duct carcinoma.

After an August 2018 protocol amendment, each cycle will be 4-week (28day) dose cycle. The first two, 4-week cycles of treatment (56 days)constitutes the dose limiting toxicity (DLT) evaluation period.Beginning with this amendment, up to 10 patients will be enrolled forevaluation at 20.0 mg/m² under the new 4-week cycle dose regimen. Ifthat dose is cleared by the SRC, subsequent dose groups will follow amodified 3+3 design. Three patients will be enrolled initially and iftoxicity issues arise, the SRC will review all pertinent data and maydecide to add more patients to fully evaluate a dose in question and/orlower the dose. If the dose is well tolerated and 3 patients reach theend of Cycle 2 without DLTs, the SRC may clear this dose and open thenext dose. The observation period for DLTs is 56 days, between Day 1through the end of Cycle 2 which includes the pre-dose assessmentsbefore receiving the Cycle 3 dose. A maximum of ten patients will beenrolled per dose level prior to the SRM. that will be conducted nolater than 5 days after at least the sixth patient dosed in each levelhas completed Cycle 2, Day 28 evaluations. The data available for thepatients dosed in all prior will be factored into the data review foreach applicable dose escalation SRM. Any dose level with 2 or more DLTsoccurring in 6 or fewer treated patients will be considered to exceedthe MTD and subsequent patients will be enrolled at lower dose level(s).After the second cycle, patients may continue to receive XMT-1536 untildisease progression if the drug is well-tolerated and patients continueto derive clinical benefit in the opinion of the Investigator. Aftercompletion of DES, the EXP segment of the study will be opened.

All adverse events will be graded according to the National CancerInstitute (NCI) Common Terminology Criteria version (CTCAE v4.03). Theobservation period for DLTs is 56 days, between Day 1 through end ofCycle 2 which includes the pre-dose assessments before receiving theCycle 3 dose. In general, adverse events Grade ≥3 are DLTs withmodifications for the following criteria: neutropenia hematologictoxicities, gastrointestinal toxicities, hepatic toxicities, andelectrolyte imbalances. XMT-1536-related toxicity that delays initiationof Cycle 3 by more than 2 weeks, hospitalization to treat aninfusion-related reaction, and any toxicity that prompts modification ofthe dose to be administered in Cycle 3 are also DLTs. Blood samplingwill be performed to determine plasma PK parameters of XMT-1536, itsrelease product auristatin F-HPA, and select metabolites. Testing foranti-drug antibodies (ADA) and neutralizing antibodies (nAb) will beperformed. Tumor responses will be Investigator-assessed using ResponseEvaluation Criteria in Solid Tumors (RECIST), version 1.1, at the end ofCycle 2 and every 2 cycles thereafter.

Study Visits

Study visits will occur. The last day of the current cycle can be thesame day as the start of the next cycle throughout the study providedthe results from the End of Cycle safety assessments are obtained andreviewed prior to initiating dosing in the next cycle. The Investigatorcan make an accommodation in the visit schedule based on a patient'smedical needs. Discuss potential schedule alterations with the MedicalMonitor prior to implementing any adjustment unless it is an emergentneed. The Investigator will evaluate every patient prior to initiating adose of XMT-1536 as described in the Schedule of Assessments. A fullphysical examination (PE) consists of examininghead/ears/eyes/nose/throat (HEENT), palpable tumors, neurological andmuscular systems, pulmonary and cardiovascular systems, abdomen andlower extremities. A brief physical examination consists of examining:HEENT, pulmonary and cardiovascular systems. Patients will be evaluatedfor the occurrence or new and/or worsening toxicities at every visit.All adverse events will be graded according to the National CancerInstitute (NCI) Common Terminology Criteria version (CTCAE v4.03).

Number of Subjects

The exact number of patients to be enrolled in DES cannot be knownprospectively and is dependent upon when the MTD or RP2D is attained.Based on preclinical animal studies, it is reasonable to expect a rangeof 21 to 40 patients to be dosed in DES. The EXP segment of the study isplanned to treat up to 30 patients each in 3 disease cohorts. Givendrop-out rates, it is reasonable to expect a range of 90 to 105 patientswill be dosed in EXP.

Eligibility

Patients who are candidates for enrollment into the study are evaluatedfor eligibility by the Investigator to ensure that the inclusion andexclusion criteria have been satisfied and that the patient is eligiblefor participation.

TABLE 1 Inclusion Criteria for DES and EXP No. Eligibility Criteria 1Females and males, age ≥18 years old. 2 ECOG performance status 0 or 1.3 Measurable disease as per RECIST, version 1.1. 4 Resolution of allacute toxic effects of prior therapy or surgical procedures to Grade ≤1(except alopecia). 5 Cardiac left ventricular ejection fraction (LVEF)≥50% or the institution’s lower limit of normal by either Echo or MUGAscan. 6 Adequate organ function as defined by the following criteria:absolute neutrophil count (ANC) ≥1500 cells/mm³, platelet count ≥100,000cells/mm³, hemoglobin ≥9 g/dL, INR, activated partial thromboplastintime (aPTT), and prothrombin time (PT) all within the institutionalupper limit of normal (ULN), serum creatinine ≤1.5 mg/dL or calculatedcreatinine clearance ≥60 mL/min¹, total bilirubin ≤ ULN, aspartateaminotransferase (AST or SGOT) and alanine aminotransferase (ALT orSGPT) ≤1.5 times the institutional upper limit of normal (ULN).albumin≥3.0 g/dL. ¹Calculated by Cockroft and Gault method. Creatinineclearance (mL/min) = (140-age) × weight (kg)/72 × (serum creatinine inmg/dL) = mL/min (for females, multiply results by 0.85). 7 Confirmedavailability (prior to Cycle 1, Day 1) of tumor tissue blocks (stronglyrecommended) or freshly cut tissue slides (See Lab Manual) for NaPi2btesting and exploratory assessments. Tissue specimens must be submittedwithin 45 days after the first dose of study drug. 8 For women ofchildbearing potential and men with partners of childbearing potential,agreement to use a highly effective form of hormonal contraception ortwo effective forms of non-hormonal contraception by the patient and/orpartner, and to continue the use of contraception for the duration ofstudy treatment and for at least 6 months after the last dose of studytreatment. Male patients whose partners are pregnant should use condomsfor the duration of the pregnancy. 9 Able to sign informed consent.

Prophylactic transfusion of blood (or blood components) prior to initialdosing cannot be used to meet enrollment criteria. Transfusion of blood(or blood components) to manage treatment-emergent anemia or othercytopenias is permissible and should be recorded as a concomitantmedication. Growth factor prophylaxis cannot be used prior to XMT-1536administration in any cycle.

TABLE 2 Disease specific inclusion criteria for DES No. EligibilityCriteria 1 Histologically or cytologically confirmed solid tumors of thetypes specified below, with incurable, locally advanced or metastaticdisease that has failed standard therapy or for which no standardtreatment option exists: Platinum-resistant ovarian cancer (includingepithelial ovarian cancer, such as, high grade serous ovarian cancer,fallopian tube, and primary peritoneal cancer), Nonsquamous NSCLC,Papillary thyroid carcinoma, Endometrial carcinoma (excludingcarcinosarcoma and stromal tumors), Papillary renal cell carcinoma,Salivary duct carcinoma.

TABLE 3 Disease specific inclusion criteria for expansion segment (EXP)No. Eligibility Criteria Cohort 1: Platinum-resistant ovarian cancer 1Histological diagnosis of high-grade serous ovarian, epithelial ovarian,fallopian tube, or primary peritoneal cancer, excluding the mucinoussubtype. 2 Platinum resistance, defined as disease progression within 6months of completing a platinum-containing chemotherapy regimen. 3 Nomore than 3 lines of prior therapy Cohort 2: Nonsquamous NSCLC 1Adenocarcinoma histological diagnosis of nonsquamous NSCLC 2 Priortreatment with a platinum-based (cisplatin or carboplatin) regimen and aPD- 1 or PD-L1 monoclonal antibody (either in combination orsequentially). 3 Patients with known oncogenic mutations for which thereare approved therapies (e.g. ALK translocation, EGFR mutation) must havedocumented intolerance or disease progression for the approved therapiesfor their mutation. Patients must have received prior treatment with aplatinum-based regimen, but prior treatment with a PD-1 or PD-L1monoclonal antibody is not required. 4 No prior treatment with acytotoxic agent or immunotherapy. Cohort 3: Additional indications 1 Forpapillary thyroid carcinoma patients, the following are required: (a)progressive, radioactive iodine-refractory, loco-regional recurrent ormetastatic disease and (b) resistance or intolerance to prior kinaseinhibitor therapy (e.g., lenvatinib, sorafenib). A patient who isconsidered inappropriate for kinase inhibitor therapy may be enrolledwith approval of the Medical Monitor. 2 For endometrial carcinomapatients, the following are required: (a) a diagnosis of epithelialendometrial carcinoma is required. Stromal tumors and carcinosarcoma(mixed malignant Mullerian tumor) are excluded and (b) a patient musthave received at least one prior chemotherapeutic regimen forendometrial cancer with carboplatin/paclitaxel or a similar regimen.Patients should have received prior hormonal therapy for endometrialcancer if appropriate, e.g., for low-grade, hormone receptor-positiveendometroid adenocarcinoma. 3 For papillary renal cell carcinomapatients, the following are required: (a) documented local confirmationof renal cell carcinoma with a predominantly papillary growth patternand (b) Progression after standard systemic therapy. 4 For salivary ductcarcinoma patients, the following are required: (a) a histologicdiagnosis of salivary duct carcinoma (other subtypes of salivary glandcancer are excluded) and (b) progression after standard systemic therapyor a lack of available effective therapy, in the assessment of theinvestigator.

TABLE 4 Exclusion Criteria for Dose Escalation and Expansion No.Eligibility Criteria Exclusion Criteria for Dose Escalation andExpansion  1 Major surgery within 28 days of starting study treatment;-or- systemic anti-cancer therapy within the lesser of 28 days or 5half-lives of the prior therapy before starting study treatment (14 daysor 5 half-lives for small molecule targeted therapy); -or- recentradiation therapy with unresolved toxicity or within a time window ofpotential toxicity (consultation with Medical Monitor is recommended). 2 Brain metastases that: are untreated, are progressive, or haverequired any type of major treatment, e.g., whole brain radiationtreatment, adjuvant chemotherapy, gamma knife, to control symptoms frombrain metastases within 30 days of the first study treatment, or anyhistory of leptomeningeal metastasis.  3 Current known active infectionwith HIV, hepatitis B virus (HBV), or hepatitis C virus (HCV). Inaddition, negative serology is required during screening for HBV andHCV: a. HBV: Patients must be negative for hepatitis B surface antigen(HBsAg) and hepatitis B core antibody (anti-HBc). Patients with evidenceof prior HBV infection (positive anti-HBc and negative HBsAg) must benegative for HBV DNA to be eligible. b. HCV: Patients must be negativefor HCV antibody, or if HCV antibody is positive, patients must have anegative PCR test for HCV RNA.  4 Current severe, uncontrolled systemicdisease (e.g., clinically significant cardiovascular, pulmonary, ormetabolic disease) or intercurrent illness that could interfere withper- protocol evaluations.  5 History of cirrhosis, hepatic fibrosis,varices, or other clinically significant liver disease. Fibroscantesting may be required for patients with a history of chronic liverdisease, e.g., fatty liver. Patients with a regular alcohol intake ofmore than one drink per day for women or more than two drinks today formen are not eligible, and alcohol consumption during trial participationshould be discouraged  6 Patients cannot receive drugs associated withhepatotoxicity concurrent with XMT-1536 administration or for 21 daysafter the last dose of XMT-1536. Patients may receiveacetaminophen/paracetamol for a limited time but at a total daily doseof ≤2 g per day. Use of NSAIDs or steroids for treatment of fever isencouraged.  7 Severe dyspnea at rest due to complications of advancedmalignancy, or requiring supplementary oxygen therapy.  8 Currentlyactive pneumonitis or interstitial lung disease  9 Pregnant or nursingwomen. 10 Diagnosis of a second malignancy that is likely to requiresystemic anticancer treatment. 11 Active corneal or conjunctivaldisease, or history of corneal or conjunctival disease within 12 monthsprior to enrollment. 12 Use of strong CYP450 inhibitors. ExclusionCriterion for Expansion 10 Use of strong CYP450 inhibitors.Use of Drugs Associated with Hepatotoxicity

Drugs Categorized by the FDA of those drugs most likely to possiblycause drug-induced liver injury (DILI) should not be used during studyparticipation. An exception for the use of paracetamol (acetaminophen)is described in Exclusion Criterion 6. If a drug on the FDA list must beused during the study to render appropriate medical care and for whichno alternative is available the Medical Monitor needs to discuss thecircumstance.

Subject Withdrawal Criteria

A patient is free to withdraw from the study at any time.

Patients who wish to terminate study participation early will berequested to continue general medical follow-up through at least 30 dayspost-dose for safety monitoring purposes, if possible. Entrance intohospice care preempts this request. Procedures described for the End ofTreatment visit will be followed.

In all cases, the reason for withdrawal must be recorded in the eCRF. Ifthe reason is initially unknown, the patient should be followed toestablish whether the reason was an adverse event. If yes, this eventwill be recorded as the reason for study termination.

Dose and Administration

XMT-1536 is provided as a colorless to yellow or brown liquid in a 5 mLround, flint glass tubing vial with a gray chlorobutyl rubber stopperwith a barrier film covered by a 20 mm aluminum, green flip-off cap.Each single-use vial contains 2.5 mL of XMT-1536 antibody drug conjugateat a concentration of 10 mg/mL and pH of 4.0 to 6.0.

Vials must be stored at −20° C. (±5° C.) in a secure,temperature-controlled freezer. XMT-1536 vials are inventoried andcontrolled using the institution's standard pharmacy procedures forcontrol of a research substance. The assigned CRA will review storageconditions and the maintenance of these conditions during periodicon-site visits.

Each XMT-1536 individual vial will be allowed to thaw at roomtemperature for up to 60 min before preparing a patient's dose. Inspectthawed vials before use: the liquid should be clear to yellow to brownwith essentially no visible particulates. Vials must not be shaken orplaced in direct sunlight. Once the dose for infusion has been prepared,it can be retained at room temperature, under typical room lightingconditions, for a maximum of 4 hours before administration.

XMT-1536 will be administered according to body surface area (BSA). TheBSA adjusted dose is calculated following each institution's standardpractice. When possible, the Mosteller Formula will be used. Thestarting dose was calculated based on height and weight collected within14 days of the first dose (can be collected the day of first dose). Doseadjustments based on subsequent weight measurements are made in accordwith each institution's standard practice. If a standard practice doesnot exist, additional weight measurements are obtained and BSA confirmedor altered prior to dosing at Cycle 2 and every two cycles thereafter.

Each dose is prepared in a 100 mL, 0.9% NS PVC infusion bag. Dosepreparation is performed according to Investigational Site procedures.Refer to Pharmacy Manual for detailed instructions. Dose preparationwill be documented in each patient's study participation record andfollow all institutional practices including quality assurance checks

The planned starting dose in the dose escalation part of the study is 3mg/m² of XMT-1536 ADC. The dose escalation scheme is shown in Table 5below. As of August 2018, DL 5 (30 mg/m2) had been cleared by SRC.However, based on emerging safety data, the SRC decided to extend theDLT observation period to two, four week cycles (56 days), amend theinclusion and exclusion criteria, and add additional patients at DL 4(20 mg/m²), with the intention to resume dose escalation per protocolguidelines if this dose is found to be tolerable. The SRC may decide to:(1) Escalate the dose to the next planned level or a higher dose levelthat is less than the planned increase based on the data demonstratingdrug tolerability. (2) Add up to 3 more patients to a the current doselevel for additional evaluation (3) De-escalate to a lower dose levelfor further evaluation; either a previously stipulated dose level or newlower dose level may be chosen based on patient safety data. (4) MTD hasbeen met and no further dose escalation will occur. At least 6 patientswill be treated at the MTD or RP2D before the expansion phase of thestudy is opened. The SRC may decide to escalate by a smaller incrementthan shown in Table 5, depending on tolerability of a given dose leveland after review of all pertinent patient data for any or all doselevels.

TABLE 5 Dose Escalation Levels Dose Escalation Start 100% 100% 67% 50%20% 20% Dose Level 1 2 3 44A 55A 66A 7A (DL) Initial number 1 1 3 3 3 33 of patients Planned dose, 3 6 12 20 30 40 (DL 6) 43 mg/m² 36 (DL 6A)

The BSA adjusted dose is calculated following each institution'sstandard practice. When possible, the Mosteller Formula is used. Thestarting dose is calculated based on height and weight collected within14 days of the first dose and can be collected on the day of first dose.Dose adjustments based on subsequent weight measurements are made inaccord with each institution's standard practice. If a standard practicedoes not exist, additional weight measurements are obtained and BSAconfirmed or altered prior to dosing at Cycle 2 and every two cyclesthereafter.

Customary supportive care medications may be used for treatment ofconcurrent, acute conditions that are associated with dosing, e.g.,hypersensitivity reaction, emesis, diarrhea and fever. Prophylactictreatments for infusion reactions or hypersensitivity are notrecommended before any patient's first dose but may be administered inadvance of the second and/or subsequent doses if these reactions occurafter the first dose. Treatment with anti-emetic and anti-pyreticmedications may be indicated for several days after infusion, and may beused before the first dose if indicated by the patient's medicalhistory. Dose adjustments after Cycle 1 are permitted under specificcircumstances.

The initial dose for each patient is administered over 90 min. If noinfusion-related reaction (IRR) occurs, all subsequent doses areadministered over 30 to 90 minutes at the discretion of theInvestigator. Treatment of IRRs should be according to institutionalstandards or the study guidelines.

Once a patient completes Cycle 1 (DLT evaluation period) doseadjustments may be made if a patient experiences toxicity but wouldbenefit from further treatment with XMT-1536, in the opinion of theinvestigator.

Criteria for Continued Dosing

In Cycle 2 and beyond, the Investigator or designee should review End ofCycle/pre-dose clinical and laboratory findings before proceeding withstudy drug administration. All liver enzymes should have recovered toGrade 1 or lower, or to the patient's baseline, before proceeding.Laboratory data, particularly liver tests, should be compared to thepatient's baseline, and delay of dosing (e.g. for 1 week) should beconsidered to allow for more complete recovery of liver function or toallow resolution of other AEs, e.g. nausea, vomiting, anorexia orfatigue.

Intrapatient Dose Reduction

Once a patient completes Cycle 1, dose adjustments may be made if apatient experiences toxicity but would benefit from further treatmentwith XMT-1536, in the opinion of the investigator. Treatment at areduced dose may commence only if the observed toxicity resolves to Gr≤1or baseline within 21 days or 28 days after the toxicity began, and atthe discretion of the Investigator after discussion with the MedicalMonitor (Table 6).

The first dose reduction will be to one dose lower than the dose levelwhere the toxicity was observed.

The second reduction will be two doses lower than the original dose atwhich the toxicity was observed.

If a third dose reduction is necessary the patient will be discontinued.

TABLE 6 Dose Reduction after Completion of Cycle 1 Dose Reduction Dose¹1^(st) One dose level lower than original dose 2^(nd) Two dose levelslower than original dose 3^(rd) Patient Discontinuation ¹The reduceddose cannot be lower than 3 mg/m² (DL 1)

In Cycle 2 or subsequent cycles, peripheral motor or sensory neuropathyGrade ≥3 that does not resolve to Grade ≤2 within 21 days afteroccurrence requires dose reduction or termination of treatment. Ingeneral, before making any dose adjustments, call the Medical Monitor todiscuss.

In Cycle 2 and beyond, the Investigator or designee should review End ofCycle/pre-dose clinical and laboratory findings before proceeding withstudy drug administration. All liver enzymes should have recovered toGrade 1 or lower, or to the patient's baseline, before proceeding.Laboratory data, particularly liver tests, should be compared to thepatient's baseline, and delay of dosing (e.g. for 1 week) should beconsidered to allow for more complete recovery of liver function or toallow resolution of other AEs, e.g. nausea, vomiting, anorexia orfatigue.

Dose Reductions in DES and EXP Due to Hepatoxicity

Patients in both DES and EXP who experience hepatic transaminase or ALPlevels of Grade ≥3 will return to office, be retested within 48 to 72hours of the initial Grade ≥3 value, and monitored at least weekly untilthe levels return to Grade ≤1 or to that patient's baseline. If thepatient resides a significant distance from the research facility suchthat return is a hardship, arrangements can be made to have a localblood draw with the results and local normal range returned to the StudyInvestigator for monitoring.

A patient who experiences one increase in hepatic transaminase or ALP toGrade ≥3 and requires longer than 72 hours to recover to Grade ≤2 (orbaseline) may continue dosing in the next cycle but at the next lowerdose.

If the patient experiences a second increase in hepatic transaminase orALP to Grade ≥3 that requires longer than 72 hours to recover to Grade≤2 (or baseline) after one prior dose reduction, he or she may continuedosing in the next cycle but at the next lower dose than that at whichthe second elevation occurred.

If the patient experiences a third Grade ≥3 elevation in hepatictransaminase or ALP, that requires longer than 72 hours to recover toGrade ≤2 (or baseline), after two prior dose reductions, he or she willbe discontinued from the study and monitored at least weekly until theselevels return to Grade ≤1 (or baseline).

XMT-1536 dose will not be modified for an increase in hepatictransaminase or ALP to Grade ≥3 if fewer than 72 hours are required torecover to Grade ≤2 (or baseline).

Any patient who experiences a Gr 4 hepatic transaminase or ALP willdiscontinue from the study and not undergo dose reduction. The Gr 4toxicity will be monitored at least weekly until return to Gr ≤1 orbaseline.

Whether or not the threshold for dose reduction is met, any increase inhepatic transaminase or ALP above Grade 1 must return to Grade ≤1 orbaseline before the next dose of study drug.

Duration of Treatment

Treatment will continue indefinitely unless one of the following occurs:disease progression, inter-current illness that prevents furtheradministration of treatment, unacceptable adverse events or pregnancy,patient non-compliance with the protocol, patient withdraws consent forparticipation from the study, and/or general or specific changes in thepatient's condition render the patient unacceptable for furthertreatment in the judgment of the Investigator.

However, should disease progression occur yet the Investigator perceivesthe potential for the patient to derive benefit from continued exposureto XMT-1536, the Investigator has the option to continue dosing afterdiscussion with the Medical Monitor.

Primary Objectives

The primary objectives in the Dose Escalation (DES) study is todetermine the maximum tolerated dose (MTD) and recommended Phase 2 dose(RP2D) of XMT-1536 administered intravenously once every three or fourweeks and assess the safety and tolerability of XMT-1536.

The primary objectives in the expansion study is to assess further thesafety and tolerability of XMT-1536 administered at the MTD/RP2Didentified in the DES, assess the preliminary anti-neoplastic activityof XMT-1536

Secondary Objectives

The secondary objective in the Dose Escalation (DES) study is to assessthe preliminary anti-neoplastic activity of XMT-1536.

The secondary objectives in the DES and expansion studies are to assessthe pharmacokinetics (PK) of XMT-1536, its release product, and selectedmetabolites, assess the development of anti-drug antibodies to XMT-1536,and assess the association of tumor expression of NaPi2b and objectivetumor response to XMT-1536.

Exploratory objectives in the DES and expansion studies are toretrospectively evaluate the association of objective response withtumor expression of genes other than NaPi2b, or other tumor molecularfeatures

Measurement of Efficacy

In both segments of the study assessments with computerized tomography(CT) scans and RECIST version 1.1 criteria are performed at the end ofCycle 2 and at the end of every even-numbered cycle thereafter. Patientsare continuously evaluated for adverse events, the use of concomitantmedications, and the occurrence of infusion reactions in any cycle.Pharmacokinetic profiles of XMT-1536 and select releaseproducts/metabolites and the development of anti-drug antibodies areevaluated. RECIST version 1.1, is used to measure tumor response. Theobjective response rate (ORR— rate of complete response [CR] and partialresponse [PR]) and disease control rate (DCR—rate of CR, PR, and stabledisease [SD]) will be measured for each cohort. Response duration,progression-free survival (PFS) and overall survival (OS) is estimated.

The primary analysis of efficacy is performed using both DES and EXPdata. The primary endpoint is ORR by Investigator radiologic review, andis defined as the proportion of patients who achieve a confirmed PR orCR per RECIST 1.1. A secondary endpoint is the DCR which is defined asthe proportion of patients who achieve complete response, partialresponse, and/or stable disease of any duration per RECIST 1.1. Thenumber and percentage of patients achieving response or clinical benefitis summarized and an exact 95% confidence interval (CI) is provided.

Analysis of other efficacy endpoints including duration of response(DOR), progression-free survival (PFS) and overall survival (OS) are bereported, where ever possible according to standard response criteria.Kaplan-Meier estimates of medians and quartiles with 95% CIs arereported for these statistics.

The efficacy endpoints are analyzed using both the efficacy analysis setand the efficacy evaluable analysis set in each cancer type cohort.

Pharmacokinetics Analysis

The PK profile of the active ingredient of XMT-1536, its release productand selected metabolites are determined for each patient bynon-compartmental analysis using standard PK software (e.g., PhoenixWinNonlin). PK parameters per patient include time of maximum observedconcentration (t_(max)), maximum concentration (C_(max)), and area underthe concentration curve for the last measurable concentration(AUC_(0-last)). When the terminal elimination phase is identified,additional parameters such as elimination half-life, clearance, andvolume of distribution are determined.

The handling of missing concentration and covariate data, outliers andvalues below the limit of quantification as well as details of themodeling for dose-response and PK parameter-response relationships areprovided in the PK analysis plan.

Statistical Analysis

A Statistical Analysis Plan (SAP) is written and contain three sections:analysis of data (1) from DES, (2) from EXP, and (3) and combined datafrom DES and EXP. This SAP will address the analysis of data recorded inthe clinical database as well as laboratory data and other data. Eachsection will be finalized before enrollment in that section hascompleted and prior to locking the database for the specific studysegment, i.e., DES or EXP. Selected tables, listings or figures(collectively referred to as TLFs) after the end of DES and EXP will begenerated for review purposes, and will be indicated in the SAP, but noanalysis report will be prepared for these intermediate reports. Onefinal analysis report will be created and included in the final ClinicalStudy. The final TLFs and clinical study report will be prepared afterdata from both phases have been locked. Any changes made to the SAP fora section after enrollment in that section has been completed will beshared with the Investigators. Any deviations from the planned analysiswill be described in the final study report.

Clinical data to PK vendor will be transferred to support PK analysis. Aseparate analysis plan addressing the PK profile of XMT-1536, itsrelease product and metabolites will be prepared.

Data from DES will be adequately summarized and reviewed with the SRC,at a minimum, before EXP dosing begins. Based on review of DES data, aprotocol amendment may be created and submitted for approval prior toinitiating the EXP segment. Statistical analyses will be carried out byNovella Clinical using SAS Version 9.3 or higher.

Continuous variables, including baseline characteristics, will besummarized by reporting the number of observations, mean, standarddeviation, median, minimum and maximum. Categorical/discrete variableswill be summarized using frequency tables showing the number andpercentage of patients within a category. Time-to-event data will besummarized using the Kaplan-Meier method.

Unless indicated otherwise, summary statistics will be reported forobserved data only. Missing data will not be imputed. If a baselinevalue is missing, no change from baseline will be calculated. Baselineis defined as the last available observation prior to the firstadministration of study drug on Cycle 1, Day 1. The handling of missingdata will be specified in the SAP along with the methods used forreporting the end points.

Results

As of February 2019, twenty patients have been dosed across 6 doselevels (3 mg/m² to 40 mg/m²), 21-day cycle and sixteen patients across 2dose levels, (20 mg/m² to 30 mg/m²), 28-day cycle. Thirteen of 20patients completed the 21-day DLT evaluation period. Sixteen of 16patients completed the 28-day evaluation period. One DLT occurred in thepatient dosed at 40 mg/m², 21-day cycle attributed to study drug and oneDLT occurred in the patient dosed at 30 mg/m², 28-day cycle, attributedto study drug. Two SAEs have occurred that were related study drug:congestive heart failure in DL 4, 2 mg/m², 21-day cycle and pyrexia inDL 5, 30 mg/m², 21-day cycle. Treatment-related adverse events (TRAEs)have been Grade 1 or 2; the most common TRAEs to date were anemia,arthralgias, fatigue, vomiting, anorexia, AST increase, diarrhea,dyspnea (unrelated or unlikely related), hypoalbuminema (unrelated orunlikely related), hypoxia (unrelated), headache, nausea, proteinuriaand vomiting. Ten patients have had post-treatment restaging scans, withstable disease (SD) as a best response for 9 patients.

As of August, 2019, fifty-one patients have been dosed across 9 doselevels (3 mg/m2 to 40 mg/m² in a 21-day regimen (n=20) and 20 mg/m²to 36mg/m² in a 28-day regimen (n=31)). One DLT occurred in one patient dosedat 30 mg/m², 28-day cycle and one DLT occurred in one patient dosed at36 mg/m², 28-day cycle. Both were considered related to XMT-1536.

Table 7 gives the characterization of the patients enrolled across 6dose levels (2 to 40 mg/m²), 21-day cycle and 2 dose levels 20 to 30mg/m²), 28-day cycle.

TABLE 7 Cycle Tumor Age Duration on Dose Dose Durations Types RangeStudy Level (mg/m²) (Days) N Y/o Min/Max 1  3 21 N = 1 55 4 cyclesOvarian 2  6 21 N = 1 54 2 cycles Ovarian 3 12 21 N = 7 39 to 67 1 to 4cycles Ovarian-1 Mean = 54 NSCLC-2 Median = 70 Endometrial-3 PapillaryRenal-1 4 20 21 N = 6 58 to 94 1 to 10 Ovarian-2 Mean = 72 cyclesNSCLC-1 Median = 70 Endometrial-1 Fallopian Tube-1 Salivary Duct-1 5 3021 N = 4 63 to 68 1 to 4 cycles Ovarian-3 Mean = 66 NSCLC-1 Median = 666 40 21 N = 1 54 1 cycle  Ovarian Enrollment Complete 4A 20 28 N = 847-74 1 to 6 cycles Ovarian Mean = 62 N = 1 Median = 63 Papillary Renal5A 30 28 N = 7 51-76 2 to 3 cycles Mean = 68 Median = 71

The protocol was amended in August, 2018. The 21-day cycle was extendedto 28 days and the observation period for DLT evaluation was extended toinclude two, 28-day cycles. This is the period after which the SafetyReview Committee evaluates patients' responses to XMT-1536 and decidesif dose escalation should occur. Table 8 is a summary of the patientsdosed as of February 2019.

TABLE 8 Dose Dosing Number of Level (DL) Regimen Patients DL 1, 3 mg/m²21-day cycle, DLT 1 observation period 1 cycle DL 2, 6 mg/m² 21-daycycle, DLT 1 observation period 1 cycle DL 3, 12 mg/m² 21-day cycle, DLT7 observation period 1 cycle DL 4, 20 mg/m² 21-day cycle, DLT 6observation period 1 cycle DL 5, 30 mg/m² 21-day cycle, DLT 4observation period 1 cycle DL 6, 40 mg/m² 21-day cycle, DLT 1observation period 1 cycle DL 4A, 20 mg/m² 28-day cycle, DLT 9observation period 2 cycle DL 5A, 30 mg/m² 28-day cycle, DLT 7observation period 2 cycle

FIG. 1 is a “swimmers plot” that summarizes the details for the time onstudy for twenty patients though DL6, 21-day cycle (n=19) and fourpatients dosed in the 28-day cycle (n=4). The plot maps patients by doselevel and tumor type on the y axis, against time on study drug on the xaxis. The x axis is organized by week time intervals. At lower doses themajority of patients came off study because of progressive disease, butstarting at DL4, most patients had stable disease.

FIG. 2 is a “waterfall plot” that shows the best response by RECISTcriteria for the patients dosed in the 21-day cycle and who have had atleast one scan to date. They axis shows change in tumor size by percent.The x-axis organizes the response by dose level. Although the number ofpatients at each dose is small, there appears to be a dose response.Starting at DL 5, two, ongoing patients reached nearly partial remission(defined as greater than or equal to a 30% reduction in tumor size frombaseline).

FIG. 3 shows the overall best response for 11 patients as well as the Hscore and CA-125 levels for the ovarian cancer patients dosed in the21-day cycle.

FIG. 4 is a “waterfall plot” that shows the best response by RECISTcriteria for the patients dosed in the 21-day cycle and 28-day cycle andwho have had at least one scan to date. The y axis shows change in tumorsize by percent. The x-axis organizes the response by dose level.Although the number of patients at each dose is small, there appears tobe a dose response. Starting at DL 5, two patients reached nearlypartial remission (defined as greater than or equal to a 30% reductionin tumor size from baseline). At DL 4A, one patient reached partialremission.

FIG. 5 is a “swimmers plot” that summarizes the details for the time onstudy for patients though DL6, 21-day cycle (n=20) and DL 4A and DL 5Apatients dosed in the 28-day cycle (n=16). The plot maps patients bydose level and tumor type on the y axis, against time on study drug onthe x axis. The x axis is organized by week time intervals. At lowerdoses the majority of patients came off study because of progressivedisease, but starting at DL4, most patients had stable disease.

As of May 10, 2019, thirty-seven patients have been dosed across 6 doselevels (3 mg/m² to 40 mg/m²), 21-day cycle and seventeen patients across2 dose levels, (20 mg/m² to 30 mg/m²), 28-day cycle. Table 9 summarizesthe patient characteristics. The treatment was well tolerated with mostadverse events being Grade 1 or 2 and no Grade 4 or 5 TRAE beingreported. Low rate of toxicities often associated withmicrotubule-targeting agents or ADCs, such as, for example, neutropenia,ocular toxicities, or peripheral neuropathy was observed.

Table 10 summarizes the TRAE in ≥10% of the patients. Thirteen patients(33%) serious adverse events (SAE) were reported. Two possible treatmentrelated SAEs were Grade 2 pyrexia at DL5 (30 mg/m²) and Grade 3 cardiacfailure congestion at DL4 (20 mg/m²), Seventeen SAEs unrelated orunlikely related to treatment occurred in 11 patients: intestinal/smallintestinal obstruction (3), disease progression (2), hypoxia (2),pleural effusion (2), abdominal pain, acute blood loss anemia,cellulitis staphylococcal, cerebrovascular accident, hemorrhagic anemia,malignant ascites, pericardial effusion, subdural hematoma, and vaginalhemorrhage. Two dose limiting toxicities were reported at DL 5A (30mg/m²) and DL 6 (40 mg/m²). At DL 5A, Grade 3 AST was elevated at cycle1, day 8 and returned to Grade 2 within 7 days and to Grade 1 within 13days. The AST elevation was accompanied by Grade 1 alkaline phosphataseelevation, but no elevation of ALT or bilirubin was reported. As DL6,Grade 3 AST was elevated at cycle 1, day 8 and returned to Grade 1within 21 days. The AST elevation was accompanied by Grade 2 ALTelevation which resolved to Grade 1 within 8 days, and Grade 1 alkalinephosphatase elevation but no elevation of bilirubin was reported.

TABLE 9 Patient Characteristics (N = 37) Age (years) Median (range) 64(39-93) Sex-N (%) Female 32 (86) Male 5 (14) ECOG performance status-N 011 (30) (%) 1 26 (70) Tumor type-N (%) Ovarian, fallopian tube, 22 (59)or primary peritoneal NSCLC 4 (11) Endometrial 8 (22) Papillary renal 2(5) Salivary duct 1 (3) Prior lines of therapy for Median (range) 4(1-13) metastatic disease Prior lines of therapy, ovarian Median (range)5 (1-11) cancer only (N = 22)

TABLE 10 Preferred Term Grade 1 Grade 2 Grade 3 Total Nausea 12 (32) 2(5) 0 14 (38) Fatigue 4 (11) 7 (19) 0 11 (30) Headache 5 (14) 5 (14) 010 (27) Aspartate 3 (8) 2 (5) 4 (11) 9 (24) aminotransferase (AST)increased Decreased appetite 1 (3) 6 (16) 0 7 (19) Blood alkaline 6 (16)0 0 6 (16) phosphatase increased Vomiting 4 (11) 1 (3) 0 5 (14) Gamma- 3(8) 1 (3) 0 4 (11) glutamyltransferase (GGT) increased Myalgia 3 (8) 1(3) 0 4 (11) Pyrexia 3 (8) 1 (3) 0 4 (10)

The plasma PK profile for the analytes (total antibody; conjugated drug(AF-HPA); free drug (AF-HPA) and the drug metabolite auristatin F (AF))was determined in blood samples from 30 patients (3 mg/m², n=1; 6 mg/m²,n=1; 12 mg/m², n=7; 20 mg/m², n=15; 30 mg/m², n=5; 40 mg/m², n=1). Theresults normalized to dose, showed that the exposure increased withincreasing doses and was nearly dose proportional. The PKcharacteristics is consistent with that reported for other approvedantibody-drug conjugates or those currently in clinical development.There was low systemic expose of free drug (AF-HPA) or its metabolite(AF) compared to that of the conjugated drug. Additionally, there was noaccumulation of free drug or its metabolite after administration ofmultiple doses.

Cut slides or whole tissue blocks from patient tumors, if available,were submitted for IHC expression analysis. Immunohistochemistry wasestablished using an automated TechMate 500 or TechMate 1000 (BioTekSolutions/Ventana medical Systems) platform, where various antigenretrieval conditions and primary antibody titrations were tested todevelop a higher through-put protocol. A protocol was selected based onstaining of control material and staining of preclinical material withknown response to ADC treatment. Briefly, for the established protocol,4μ sections were cut, dewaxed, and rehydrated through xylene and aseries of alcohols. Slides were antigen retrieved in a standard steamerusing BioGenex AR-10 retrieval solution. On the TechMate platform,further retrieval was performed with Proteinase K (DAKO). Followingserum blocking: the primary antibody (anti-NaPi2b/MERS67) was appliedfor 30 minutes at room temperature, then endogenous peroxidase block andnon-biotin polymer-based detection (rabbit Polink-2 Plus detectionsystem, GBI) was used and finally a brief hematoxylin counterstaining.

Tumor membrane staining was evaluated by manual reading on a lightmicroscope using an “H-Score” method. A hematoxylin and eosin stainedslide as well as a rabbit IgG isotype control stained slide wereevaluated as part of the scoring process. The same scoring method wasused for all evaluated tumor types. The H-score incorporates stainingintensity (determined by increasing intensity from 0 to 3+) as well aspercent cells positive detected at the tumor cell membrane. H-Score=(%at <1+X 0)+((% at 1+)X 1)+((% at 2+)X 2)+((%3+) X3). Hypocellularspecimens or those where the primary tumor was not represented in thesubmitted specimen are not shown.

FIG. 6 shows the NaPi2b protein expression in 34 patient tumor typese.g., ovarian, NSCLC, endometrial, papillary renal and salivary duct)wherein the fallopian tube and primary peritoneal tumor types arecombined with the ovarian tumor types. The y axes is the H score in therange 0-300 for the number of positive tumor cells and stainingintensity in NaPi2b IHC assay. Detectable NaPi2b protein expression wasobserved in all ovarian and lung adenocarcinoma samples.

FIG. 7 is a “swimmers plot” that summarizes the details for the time onstudy for patients though DL6, 21-day cycle (n=20) and DL 4A and DL 5Apatients dosed in the 28-day cycle (n=17). The plot maps patients bydose level and tumor type on the y axis, against time on study drug onthe x axis. The x axis is organized by week time intervals. At lowerdoses the majority of patients came off study because of progressivedisease, but starting at DL4, most patients had stable disease.

Table 11 summarizes the reason for discontinuation participation in thestudy across the dose levels. Table 12 gives the outcome response forthe evaluable population. Based on objective responses and duration oftreatment, clinical activity was observed at doses of 20 mg/m² orhigher.

TABLE 11 DL1 DL2 DL3 DL4 & 4A DL5 & 5A DL6 3 mg/m² 6 mg/m² 12 mg/m² 20mg/m² 30 mg/m² 40 mg/m² Total n = 1 n = 1 n = 7 n = 13 n = 11 n = 1 N =34^(b) Progressive 1 1 3 7 6 18 (53%) Disease per RECIST Clinical 4 2 39 (26%) Progression ^(a) Patient 2 1 1 4 (12%) Choice Physician 2 1 3(9%) Choice ^(a) Death in 3 of 10 patients, none related to XMT-1536^(b)3 patients are ongoing

TABLE 12 All Ovarian OC + Cancer OC NSCLC NSCLC OC Outcome (OC) AllNSCLC ≥20 mg/m² ≥20 mg/m² ≥20 mg/m² ≥30 mg/m² N 19 3 16 2 18 7 PR* 3(16%) 0 (0%) 3 (19%) 0 (0%) 3 (17%) 2 (28%) SD* 8 (42%) 2 (67%) 6 (38%)2 (100%) 8 (44%) 3 (43%) DCR (PR + 11 (58%) 2 (67%) 9 (57%) 2 (100%) 11(61%) 5 (71%) SD) Treatment 8 (42%) 1 (33%) 9 (57%) 1 (50%) 10 (56%) 3(43%) duration >16 weeks PD* 8 (42%) 1 (33%) 7 (43%) 0 (0%) 7 (39%) 2(28%) *As measured by RECIST, version 1.1

A 70-year-old woman with platinum-resistant high-grade serous ovariancancer and 11 prior lines of therapy was treated at DL4A (20 mg/m²).Target lesions of perihepatic and mid-abdominal metastases, 52 and 42 mmrespectively, decrease 40% in diameter at the end of Cycle 2 (4-weekcycles) and 75% at the end of Cycle 3. Objective responses have beenobserved at doses of 20 mg/m² and higher. In ovarian cancer and lungadenocarcinoma patients treated at ≥20 mg/m² (N=18), 3 (17%) PRs and 8(44%) SDs have been observed (ORR 17%, SD 44%, DCR 61%), with treatmentduration of >16 weeks in 9 patients (50%) at doses of 20 mg/m² andhigher (n=18).

Table 13 gives the characterization of the patients enrolled across 7dose levels (2 to 40 mg/m²), 21-day cycle and 3 dose levels 20 to 36mg/m²), 28-day cycle as of August 2019.

TABLE 13 Dose Dose Cycle Durations Tumor Types Age Range Duration onStudy Level (mg/m²) (Days) N Y/o Min/Max 1  3 21 N = 1 55 4 cyclesOvarian 2  6 21 N = 1 54 2 cycles Ovarian 3 12 21 N = 7 39 to 67 1 to 4cycles Ovarian-1 Mean = 54 NSCLC-2 Median = 70 Endometrial-3 PapillaryRenal-1 4 20 21 N = 6 58 to 94 1 to 10 Ovarian-2 Mean = 72 cyclesNSCLC-1 Median = 70 Endometrial-1 Fallopian Tube-1 Salivary Duct-1 5 3021 N = 4 63 to 68 1 to 4 cycles Ovarian-3 Mean = 66 NSCLC-1 Median = 666 40 21 N = 1 54 1 cycle Ovarian Enrollment Complete 4A 20 28 N = 847-74 1 to 12 Ovarian Mean = 62 cycles N = 1 Median = 63 Papillary Renal5A 30 28 N = 8 51-76 2 to 6 cycles Ovarian-4 Mean = 68 Endometrial-4Median = 71 5A 30 28 N = 7 51-85 2 cycles Additional Ovarian-6 Mean = 68NSCLC-1 Median = 71 6A 28 N = 7 47-63 2 cycles Ovarian-5 Mean = 54NSCLC-2 Median = 56

The protocol was amended in July 2019. Eligibility criteria forenrolling ovarian cancer patients and non-small lung cancer patientswere added. Table 14 is a summary of the patients dosed as of August2019.

TABLE 14 Dose Dosing Number of Level (DL) Regimen Patients DL 1, 3 mg/m²21-day cycle, DLT 1 observation period 1 cycle DL 2, 6 mg/m² 21-daycycle, DLT 1 observation period 1 cycle DL 3, 12 mg/m² 21-day cycle, DLT7 observation period 1 cycle DL 4, 20 mg/m² 21-day cycle, DLT 6observation period 1 cycle DL 5, 30 mg/m² 21-day cycle, DLT 4observation period 1 cycle DL 6, 40 mg/m² 21-day cycle, DLT 1observation period 1 cycle DL 4A, 20 mg/m² 28-day cycle, DLT 9observation period 2 cycle DL 5 A, 30 mg/m² 28-day cycle, DLT 8observation period 2 cycle DL 5 A, 30 mg/m² 28-day cycle, DLT 7additional observation period 2 cycle DL 6 A, 36 mg/m² 28-day cycle, DLT7 observation period 2 cycleFIG. 8 is a “swimmers plot” that summarizes the details for the time onstudy for patients across 7 dose levels (2 to 40 mg/m²), 21-day cycleand 3 dose levels (20 to 36 mg/m²), 28 day cycle. The plot maps patientsby dose level and tumor type on the y axis, against time on study drugon the x axis. The x axis is organized by week time intervals. At lowerdoses the majority of patients came off study because of progressivedisease, but starting at DL4, most patients had stable disease. Maximumduration on study for a patient ongoing in the trial is 50 weeks.

CONCLUSIONS

XMT-1536 has been well-tolerated up to the 30 mg/m² or 36 mg/m² or 43mg/m² dose level with early signs of anti-tumor activity. Neither MTDnor RP2D has been identified. Dose escalation continues in patients withNaPi2b-expressing ovarian cancer, NSCLC, papillary thyroid cancer,endometrial cancer, papillary renal cell cancer and salivary duct cancer

OTHER EMBODIMENTS

While the invention has been described in conjunction with the detaileddescription thereof, the foregoing description is intended to illustrateand not limit the scope of the invention, which is defined by the scopeof the appended claims. Other aspects, advantages, and modifications arewithin the scope of the following claims.

What is claimed is:
 1. A method of treating a NaPi2b expressing tumor ina subject in need thereof, the method comprising administering to thesubject a NaPi2b-targeted antibody polymer-drug conjugate by infusion ata dose of about 20 mg/m² or about 30 mg/m² on the first day of treatmentand every four weeks thereafter, wherein the conjugate comprises aNaPi2b antibody comprising: a CDRH1 comprising the amino acid sequenceGYTFTGYNIH (SEQ ID NO: 5); a CDRH2 comprising the amino acid sequenceAIYPGNGDTSYKQKFRG (SEQ ID NO: 6); a CDRH3 comprising the amino acidsequence GETARATFAY (SEQ ID NO: 7); a CDRL1 comprising the amino acidsequence SASQDIGNFLN (SEQ ID NO: 8); a CDRL2 comprising the amino acidsequence YTSSLYS (SEQ ID NO: 9); and a CDRL3 comprising the amino acidsequence QQYSKLPLT (SEQ ID NO: 10); and a polymer-drug conjugate ofFormula A:

wherein: the polymer comprises poly(1-hydroxymethylethylenehydroxymethyl-formal) (PHF) having a molecular weight ranging from about5 kDa to about 10 kDa; m is an integer from 20 to 75, m₁ is an integerfrom about 5 to about 35, m₂ is an integer from about 3 to about 10,m_(3a) an integer from 0 to about 4, m_(3b) is an integer from 1 toabout 5, the sum of m, m₁, m₂, m_(3a), and m_(3b) ranges from about 40to about 75, and m₅ is an integer from about 2 to about
 5. 2. The methodof claim 1, wherein the NaPi2b antibody comprises a variable heavy chaincomprising the amino acid sequence of SEQ ID NO: 3 and a variable lightchain comprising the amino acid sequence of SEQ ID NO: 4; and a heavychain comprising the amino acid sequence of SEQ ID NO: 1 and a lightchain comprising the amino acid sequence of SEQ ID NO:
 2. 3. The methodof claim 1, wherein the subject is human.
 4. The method of claim 1,wherein the dose is about 20 mg/m².
 5. The method of claim 1, whereinthe dose is about 30 mg/m².
 6. The method of claim 1, wherein the tumoris selected from the group consisting ovarian cancer, non-small celllung cancer (NSCLC), papillary thyroid cancer, endometrial cancer,papillary renal cell cancer, cholangiocarcinoma, salivary duct cancer,clear cell renal cancer, breast cancer, kidney cancer or cervicalcancer.
 7. The method of claim 6, wherein the ovarian cancer isepithelial ovarian cancer, fallopian tube cancer, or primary peritonealcancer.
 8. The method of claim 7, wherein the epithelial ovarian canceris subtyped as high-grade serous ovarian cancer, low-grade ovariancancer or clear cell ovarian cancer.
 9. The method of claim 8, whereinthe ovarian cancer is platinum resistant and the subject has received nomore than 3 line of prior therapy.
 10. The method of claim 4, whereinthe subject has ovarian cancer and has received no more than 3 line ofprior therapy.
 11. The method of claim 6, wherein the NSCLC cancer isnon-squamous and is sub-typed as adenocarcinoma.
 12. The method of claim6, wherein the subject has NSCLC and has received prior treatment with aplatinum-based regimen and a PD-1 or PD-L1monoclonal antibody.
 13. Themethod of claim 12, wherein the platinum-based regimen and PD-1 or PD-L1monoclonal antibody are administered in combination.
 14. The method ofclaim 12, wherein the platinum-based regimen and PD-1 or PD-L1monoclonal antibody are administered sequentially.
 15. The method ofclaim 11, wherein the subject has received no treatment with a cytotoxicagent or has received no immunotherapy treatment.
 16. The method ofclaim 6, wherein the papillary thyroid cancer is progressive,radioactive iodine-refractory, loco-regional recurrent or metastaticdisease.
 17. The method of claim 16, wherein the subject has papillarythyroid cancer and is resistance or intolerance to kinase inhibitortherapy.
 18. The method of claim 6, wherein the endometrial cancer isepithelial endometrial cancer and is not a stromal tumor or acarcinosarcoma.
 19. The method of claim 6, wherein the papillary renalcell cancer has a predominantly papillary growth pattern.
 20. The methodclaims 6, wherein the cancer has a histologic diagnosis of salivary ductcancer has progressed after standard systemic therapy.